Cell Animal Models Research Articles

Efficacy of Bone Regeneration Cell Therapy Using Mesenchymal Stem Cells Originating from Embryonic Stem Cells in Animal Models; Bone Defects and Osteomyelitis.

Bone defects are commonly encountered due to accidents, diseases, or aging, and the demand for effective bone regeneration, particularly for dental implants, is increasing in our aging society. Mesenchymal stem cells (MSCs) are promising candidates for regenerative therapies; however, obtaining sufficient quantities of these cells for clinical applications remains challenging. DW-MSCs, derived from embryonic stem cells and developed by Daewoong Pharmaceutical, exhibit a robust proliferative capacity even after extensive culture. This study explores the therapeutic potential of DW-MSCs in various animal models of bone defects. DW-MSCs were expanded for over 13 passages for in vivo use in rat and canine models of bone defects and osteomyelitis. The research focused on the in vivo osteogenic differentiation of DW-MSCs, the establishment of a fibrin-based system for bone regeneration, the assessment of bone repair following treatment in animal models, and comparisons with commercially available bone grafts. Results showed that DW-MSCs exhibited superior osteogenic differentiation compared to other materials, and the fibrinization process not only preserved but enhanced their proliferation and differentiation capabilities through a 3D culture effect. In both bone defect models, DW-MSCs facilitated significant bone regeneration, reduced inflammatory responses in osteomyelitis, and achieved effective bone healing. The therapeutic outcomes of DW-MSCs were comparable to those of commercial bone grafts but demonstrated qualitatively superior bone tissue restructuring. Our findings suggest that DW-MSCs offer a promising approach for bone regeneration therapies due to their high efficacy and anti-inflammatory properties.

Role of Complement in Liver Diseases.

This review aims to summarize recent research using animal models, cell models, and human data regarding the role of complement in liver disease. Complement is part of the innate immune system and was initially characterized for its role in control of pathogens. However, evidence now indicates that complement also plays an important role in the response to cellular injury that is independent of pathogens. The liver is the main organ responsible for producing circulating complement. In response to liver injury, complement is activated and likely plays a dual role, both contributing to and protecting from injury. In uncontrolled complement activation, cell injury and liver inflammation occur, contributing to progression of liver disease. Complement activation is implicated in the pathogenesis of multiple liver diseases, including alcohol-associated liver disease, metabolic dysfunction-associated steatotic liver disease, fibrosis and cirrhosis, hepatocellular carcinoma, and autoimmune hepatitis. However, the mechanisms by which complement is overactivated in liver diseases are still being unraveled.

B cells abnormalities in asthma

Abstract. Asthma is a chronic inflammation in the respiratory tract, which is an abnormal immune response of type I allergic reaction with elevated IgE levels. The abnormality of B cell contributing to asthma development has not been elucidated completely. Recently, data from different research groups showed that both CD27+CD38+ plasmablasts and CD24hiCD38hi transitional B cells are capable to induce the production of IgE antibody. In contrast, Bregs is crucial for inhibiting type 2 inflammation through secreting cytokines IL-10 in asthma patients of both children and adults. With the advancement of high-throughput sequencing and analysis technology in recent years, there has been further displaying of the association between B subgroups and asthma besides Breg and plasmablast subsets. This article briefly summarized the pathogenesis and pathological progression of Breg cells in animal models and patients of asthma, with a particular focus on high-throughput sequencing revealing new mechanisms and diagnostic

Longitudinal multi-omics reveals pathogenic TSC2 variants disrupt developmental trajectories of human cortical organoids derived from Tuberous Sclerosis Complex.

Tuberous Sclerosis Complex (TSC), an autosomal dominant condition, is caused by heterozygous mutations in either the TSC1 or TSC2 genes, manifesting in systemic growth of benign tumors. In addition to brain lesions, neurologic sequelae represent the greatest morbidity in TSC patients. Investigations utilizing TSC1/2 -knockout animal or human stem cell models suggest that TSC deficiency-causing hyper-activation of mTOR signaling might precipitate anomalous neurodevelopmental processes. However, how the pathogenic variants of TSC1/2 genes affect the longitudinal trajectory of human brain development remains largely unexplored. Here, we employed 3-dimensional cortical organoids derived from induced pluripotent stem cells (iPSCs) from TSC patients harboring TSC2 variants, alongside organoids from age- and sex-matched healthy individuals as controls. Through comprehensively longitudinal molecular and cellular analyses of TSC organoids, we found that TSC2 pathogenic variants dysregulate neurogenesis, synaptogenesis, and gliogenesis, particularly for reactive astrogliosis. The altered developmental trajectory of TSC organoids significantly resembles the molecular signatures of neuropsychiatric disorders, including autism spectrum disorders, epilepsy, and intellectual disability. Intriguingly, single cell transcriptomic analyses on TSC organoids revealed that TSC2 pathogenic variants disrupt the neuron/reactive astrocyte crosstalk within the NLGN-NRXN signaling network. Furthermore, cellular and electrophysiological assessments of TSC cortical organoids, along with proteomic analyses of synaptosomes, demonstrated that the TSC2 variants precipitate perturbations in synaptic transmission, neuronal network activity, mitochondrial translational integrity, and neurofilament formation. Notably, similar perturbations were observed in surgically resected cortical specimens from TSC patients. Collectively, our study illustrates that disease-associated TSC2 variants disrupt the neurodevelopmental trajectories through perturbations of gene regulatory networks during early cortical development, leading to mitochondrial dysfunction, aberrant neurofilament formation, impaired synaptic formation and neuronal network activity.

AUTS2 disruption causes neuronal differentiation defects in human cerebral organoids through hyperactivation of the WNT/β-catenin pathway

Individuals with the Autism Susceptibility Candidate 2 (AUTS2) gene disruptions exhibit symptoms such as intellectual disability, microcephaly, growth retardation, and distinct skeletal and facial differences. The role of AUTS2 in neurodevelopment has been investigated using animal and embryonic stem cell models. However, the precise molecular mechanisms of how AUTS2 influences neurodevelopment, particularly in humans, are not thoroughly understood. Our study employed a 3D human cerebral organoid culture system, in combination with genetic, genomic, cellular, and molecular approaches, to investigate how AUTS2 impacts neurodevelopment through cellular signaling pathways. We used CRISPR/Cas9 technology to create AUTS2-deficient human embryonic stem cells and then generated cerebral organoids with these cells. Our transcriptomic analyses revealed that the absence of AUTS2 in cerebral organoids reduces the populations of cells committed to the neuronal lineage, resulting in an overabundance of cells with a transcription profile resembling that of choroid plexus (ChP) cells. Intriguingly, we found that AUTS2 negatively regulates the WNT/β-catenin signaling pathway, evidenced by its overactivation in AUTS2-deficient cerebral organoids and in luciferase reporter cells lacking AUTS2. Importantly, treating the AUTS2-deficient cerebral organoids with a WNT inhibitor reversed the overexpression of ChP genes and increased the downregulated neuronal gene expression. This study offers new insights into the role of AUTS2 in neurodevelopment and suggests potential targeted therapies for neurodevelopmental disorders.

Sculpting the tumour microenvironment by combining radiotherapy and ATR inhibition for curative-intent adjuvant immunotherapy

The combination of radiotherapy/chemoradiotherapy and immune checkpoint blockade can result in poor outcomes in patients with locally advanced head and neck squamous cell carcinoma (HNSCC). Here, we show that combining ATR inhibition (ATRi) with radiotherapy (RT) increases the frequency of activated NKG2A+PD-1+ T cells in animal models of HNSCC. Compared with the ATRi/RT treatment regimen alone, the addition of simultaneous NKG2A and PD-L1 blockade to ATRi/RT, in the adjuvant, post-radiotherapy setting induces a robust antitumour response driven by higher infiltration and activation of cytotoxic T cells in the tumour microenvironment. The efficacy of this combination relies on CD40/CD40L costimulation and infiltration of activated, proliferating memory CD8+ and CD4+ T cells with persistent or new T cell receptor (TCR) signalling, respectively. We also observe increased richness in the TCR repertoire and emergence of numerous and large TCR clonotypes that cluster based on antigen specificity in response to NKG2A/PD-L1/ATRi/RT. Collectively, our data point towards potential combination approaches for the treatment of HNSCC.

A Review on in-vivo and in-vitro Models of Obesity and Obesity-Associated Co-Morbidities.

Obesity is becoming a global pandemic with pandemic proportions. According to the WHO estimates, there were over 1.9 billion overweight individuals and over 650 million obese adults in the globe in 2016. In recent years, scientists have encountered difficulties in choosing acceptable animal models, leading to a multitude of contradicting aspects and incorrect outcomes. This review comprehensively evaluates different screening models of obesity and obesity-associated comorbidities to reveal the advantages and disadvantages/limitations of each model while also mentioning the time duration each model requires to induce obesity. For this review, the authors have gone through a vast number of article sources from different scientific databases, such as Google Scholar, Web of Science, Medline, and PubMed. In-vivo models used to represent a variety of obesity-inducing processes, such as diet-induced, drug-induced, surgical, chemical, stress-induced, and genetic models, are discussed. Animal cell models are examined with an emphasis on their use in understanding the molecular causes of obesity, for which we discussed in depth the important cell lines, including 3T3-L1, OP9, 3T3-F442A, and C3H10T1/2. Screening models of obesity-associated co-morbidities like diabetes, asthma, cardiovascular disorders, cancer, and polycystic ovarian syndrome (PCOS) were discussed, which provided light on the complex interactions between obesity and numerous health problems. Mimicking obesity in an animal model reflects multifactorial aspects is a matter of challenge. Future studies could address the ethical issues surrounding the use of animals in obesity research as well as investigate newly developed models, such as non-mammalian models. In conclusion, improving our knowledge and management of obesity and related health problems will require ongoing assessment and improvement of study models.

A Dynamic Interplay of Innate Immune Responses During Urinary Tract Infection.

Urinary tract infections (UTIs) represent one of the most prevalent bacterial infections globally, manifesting in diverse clinical phenotypes with varying degrees of severity and complications. The mechanisms underlying UTIs are gradually being elucidated, leading to an enhanced understanding of the immune responses involved. Innate immune cells play a crucial defensive role against uropathogenic bacteria through various mechanisms. Despite their significant contributions to host defense, these cells often fail to achieve complete clearance of uropathogens, necessitating the frequent prescription of antibiotics for UTI patients. However, the persistence of infections and related pathological symptoms in the absence of innate immune cells in animal models underscore the importance of innate immunity in UTIs. Therefore, the host protective functions of innate immune cells, including neutrophils, macrophages, mast cells, NK cells, innate lymphoid cells, and γδ T cells, are delicately coordinated and timely regulated by a variety of cytokines to ensure successful pathogen clearance.

Evolving Role of Exosomes in Plastic and Reconstructive Surgery and Dermatology.

Exosomes, or extracellular vesicles, represent the latest cell-free addition to the regenerative medicine toolkit. In vitro preclinical studies have demonstrated the safety and efficacy of exosomes, which vary based on source and biomanufacturing, for a myriad of potential therapeutic applications relevant to skin and soft tissue reconstruction. Primary search was performed in September 2021 on the MEDLINE database via PubMed and Ovid, with focus on articles about therapeutic application of exosomes or extracellular vesicles. In total, 130 articles met criteria for applicability, including early-stage clinical trials, preclinical research studies with in vivo application, and articles applicable to plastic and reconstructive surgery and dermatology. Most studies used animal models of human disease processes, using either animal donor cells to isolate exosomes, or human donor cells in animal models. Exosome technology has catapulted as an acellular therapeutic vehicle with off-the-shelf accessibility. These features eliminate prior threshold for broad adoption of regenerative cell-based therapies into surgical and medical practice. To date, there are no exosome products approved by the US Food and Drug Administration. This review highlights exosomes as the new frontier in regenerative medicine and outlines its preclinical therapeutic applications for cutaneous repair and restoration.

CD4+ T-Cell Senescence in Neurodegenerative Disease: Pathogenesis and Potential Therapeutic Targets.

With the increasing proportion of the aging population, neurodegenerative diseases have become one of the major health issues in society. Neurodegenerative diseases (NDs), including multiple sclerosis (MS), Alzheimer's disease (AD), Parkinson's disease (PD), and amyotrophic lateral sclerosis (ALS), are characterized by progressive neurodegeneration associated with aging, leading to a gradual decline in cognitive, emotional, and motor functions in patients. The process of aging is a normal physiological process in human life and is accompanied by the aging of the immune system, which is known as immunosenescence. T-cells are an important part of the immune system, and their senescence is the main feature of immunosenescence. The appearance of senescent T-cells has been shown to potentially lead to chronic inflammation and tissue damage, with some studies indicating a direct link between T-cell senescence, inflammation, and neuronal damage. The role of these subsets with different functions in NDs is still under debate. A growing body of evidence suggests that in people with a ND, there is a prevalence of CD4+ T-cell subsets exhibiting characteristics that are linked to senescence. This underscores the significance of CD4+ T-cells in NDs. In this review, we summarize the classification and function of CD4+ T-cell subpopulations, the characteristics of CD4+ T-cell senescence, the potential roles of these cells in animal models and human studies of NDs, and therapeutic strategies targeting CD4+ T-cell senescence.

Hypoxia preconditioning increases Notch1 activity by regulating DNA methylation in vitro and in vivo.

Our previous research has demonstrated that hypoxic preconditioning (HPC) can improve spatial learning and memory abilities in adult mice. Adult hippocampal neurogenesis has been associated with learning and memory. The Neurogenic locus notch homolog protein (Notch) was involved in adult hippocampal neurogenesis, as well as in learning and memory. It is currently unclear whether the Notch pathway regulates hippocampal neuroregeneration by modifying the DNA methylation status of the Notch gene following HPC. The HPC animal model and cell model were established through repeated hypoxia exposure using mice and the mouse hippocampal neuronal cell line HT22. Step-down test was conducted on HPC mice. Real-time PCR and Western blot analysis were used to assess the mRNA and protein expression levels of Notch1 and hairy and enhancer of split1 (HES1). The presence of BrdU-positive cells and Notch1 expression in the hippocampal dental gyrus (DG) were examined with confocal microscopy. The methylation status of the Notch1 was analyzed using methylation-specific PCR (MS-PCR). HT22 cells were employed to elucidate the impact of HPC on Notch1 in vitro. HPC significantly improved the step-down test performance of mice with elevated levels of mRNA and protein expression of Notch1 and HES1 (P < 0.05). The intensities of the Notch1 signal in the control group, the H group and the HPC group were 2.62 ± 0.57 × 107, 2.87 ± 0.84 × 107, and 3.32 ± 0.14 × 107, respectively, and the number of BrdU (+) cells in the hippocampal DG were 1.83 ± 0.54, 3.71 ± 0.64, and 7.29 ± 0.68 respectively. Compared with that in C and H group, the intensity of the Notch1 signal and the number of BrdU (+) cells increased significantly in HPC group (P < 0.05). The methylation levels of the Notch1 promoter 0.82 ± 0.03, 0.65 ± 0.03, and 0.60 ± 0.02 in the C, H, and HPC groups, respectively. The methylation levels of Notch1 decreased significantly (P < 0.05). The effect of HPC on HT22 cells exhibited similarities to that observed in the hippocampus. HPC may confer neuroprotection by activating the Notch1 signaling pathway and regulating its methylation level, resulting in the regeneration of hippocampal neurons.

Disease Mechanisms and Therapeutic Approaches in SMARD1-Insights from Animal Models and Cell Models.

Spinal muscular atrophy with respiratory distress type 1 (SMARD1) is a fatal childhood motoneuron disease caused by mutations in the IGHMBP2 gene. It is characterized by muscle weakness, initially affecting the distal extremities due to the degeneration of spinal α-motoneurons, and respiratory distress, due to the paralysis of the diaphragm. Infantile forms with a severe course of the disease can be distinguished from juvenile forms with a milder course. Mutations in the IGHMBP2 gene have also been found in patients with peripheral neuropathy Charcot-Marie-Tooth type 2S (CMT2S). IGHMBP2 is an ATP-dependent 5'→3' RNA helicase thought to be involved in translational mechanisms. In recent years, several animal models representing both SMARD1 forms and CMT2S have been generated to initially study disease mechanisms. Later, the models showed very well that both stem cell therapies and the delivery of the human IGHMBP2 cDNA by AAV9 approaches (AAV9-IGHMBP2) can lead to significant improvements in disease symptoms. Therefore, the SMARD1 animal models, in addition to the cellular models, provide an inexhaustible source for obtaining knowledge of disease mechanisms, disease progression at the cellular level, and deeper insights into the development of therapies against SMARD1.

Abstract 7132: Preclinical characterization of LAE119, a novel PARP1 selective inhibitor and trapper

Abstract Current PARP inhibitors primarily target PARP1/2 and demonstrate high efficacy against tumors with deficiencies in homologous recombination repair, such as those with BRCA mutations. However, these inhibitors are also associated with significant hematological toxicities. Studies have indicated that the synthetic lethality observed with BRCA mutations is mainly driven by PARP1, while PARP2 is not essential in this context. PARP2 has been shown to be crucial for the survival of hematopoietic stem/progenitor cells in animal models. Therefore, selective inhibition of PARP1 could potentially reduce toxicity and improve therapeutic outcomes. Here we introduce LAE119, a potent and selective PARP1 inhibitor and PARP1-DNA trapper. It demonstrates more than a 1000-fold selectivity for PARP1 DNA trapping compared to PARP2. In comparison to most PARP inhibitors including AZD5305, LAE119 exhibits extremely long residence time on PARP1 in both biochemical and cellular assays. It demonstrates robust anti-proliferation activity against BRCA2−/− DLD-1 cells and MDA-MB-436 cells, with IC50 value of 0.6 nM in both cell lines. In CAL51 cells, which has a BRCA2 mutation but with low PARP1 expression level and are much less sensitive to AZD5305, LAE119 demonstrates much better activity than AZD5305, particularly when PARP1 level is increased with transgene expression. LAE119 demonstrated substantial tumor inhibition in MDA-MB-436 and Capan-1 xenograft models. LAE119 also displays favorable ADME properties and PK profiles. It has high tumor/plasma ratio which is beneficial for sustainable anti-tumor effect and reduction of off-target effects. In rat hematologic toxicity study, LAE119 at 10 mpk QD dose was well tolerated and had minimal effects on hematologic parameters. Both in vitro and in vivo data strongly support the further clinical investigation of LAE119. Citation Format: Ming Li, Yan Chen, Junyan Chen, Ling Jiang, Xiaofen Lin, Justin Gu. Preclinical characterization of LAE119, a novel PARP1 selective inhibitor and trapper [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 7132.

Interleukin-22 Alleviates Caerulein-Induced Acute Pancreatitis by Activating AKT/mTOR Pathway

BackgroundAcute pancreatitis (AP) is one of the most common acute abdominal disorders; due to the lack of specific treatment, the treatment of acute pancreatitis, especially serious acute pancreatitis (SAP), is difficult and challenging. We will observe the changes of Interleukin -22 levels in acute pancreatitis animal models, and explore the mechanism of Interleukin -22 in acute pancreatitis.ObjectiveThis study aims to assess the potential protective effect of Interleukin -22 on caerulein-induced acute pancreatitis and to explore its mechanism.MethodsBlood levels of amylase and lipase and Interleukin -22 were assessed in mice with acute pancreatitis. In animal model and cell model of caerulein-induced acute pancreatitis, the mRNA levels of P62 and Beclin-1 were determined using PCR, and the protein expression of P62, LC3-II, mTOR, AKT, p-mTOR, and p-AKT were evaluated through Western blot analysis.ResultsInterleukin -22 administration reduced blood amylase and lipase levels and mitigated tissue damage in acute pancreatitis mice model. Interleukin -22 inhibited the relative mRNA levels of P62 and Beclin-1, and the Interleukin -22 group showed a decreased protein expression of LC3-II and P62 and the phosphorylation of the AKT/mTOR pathway. Furthermore, we obtained similar results in the cell model of acute pancreatitis.ConclusionThis study suggests that Interleukin -22 administration could alleviate pancreatic damage in caerulein-induced acute pancreatitis. This effect may result from the activation of the AKT/mTOR pathway, leading to the inhibition of autophagy. Consequently, Interleukin -22 shows potential as a treatment.

Reprogramming of astrocytes to neuronal-like cells in spinal cord injury: a systematic review.

A Systematic Review OBJECTIVES: To determine the therapeutic efficacy of in vivo reprogramming of astrocytes into neuronal-like cells in animal models of spinal cord injury (SCI). PRISMA 2020 guidelines were utilized, and search engines Medline, Web of Science, Scopus, and Embase until June 2023 were used. Studies that examined the effects of converting astrocytes into neuron-like cells with any vector in all animal models were included, while conversion from other cells except for spinal astrocytes, chemical mechanisms to provide SCI models, brain injury population, and conversion without in-vivo experience were excluded. The risk of bias was calculated independently. 5302 manuscripts were initially identified and after eligibility assessment, 43 studies were included for full-text analysis. After final analysis, 13 manuscripts were included. All were graded as high-quality assessments. The transduction factors Sox2, Oct4, Klf4, fibroblast growth factor 4 (Fgf4) antibody, neurogenic differentiation 1 (Neurod1), zinc finger protein 521 (Zfp521), ginsenoside Rg1, and small molecules (LDN193189, CHIR99021, and DAPT) could effectively reprogramme astrocytes into neuron-like cells. The process was enhanced by p21-p53, or Notch signaling knockout, valproic acid, or chondroitin sulfate proteoglycan inhibitors. The type of mature neurons was both excitatory and inhibitory. Astrocyte reprogramming to neuronal-like cells in an animal model after SCI appears promising. The molecular and functional improvements after astrocyte reprogramming were demonstrated in vivo, and further investigation is required in this field.

Current Status and Prospects of Viral Vector-Based Gene Therapy to Treat Kidney Diseases.

Inherited kidney diseases are among the leading causes of chronic kidney disease, reducing the quality of life and resulting in substantial socioeconomic impact. The advent of early genetic testing and the growing understanding of the molecular basis and pathophysiology of these disorders have opened avenues for novel treatment strategies. Viral vector-based gene therapies have evolved from experimental treatments for rare diseases to potent platforms that carry the intrinsic potential to provide a cure with a single application. Several gene therapy products have reached the market, and the numbers are only expected to increase. Still, none target inherited kidney diseases. Gene transfer to the kidney has lagged when compared to other tissue-directed therapies such as hepatic, neuromuscular, and ocular tissues. Systemic delivery of genetic information to tackle kidney disease is challenging. The pharma industry is taking steps to take on kidney disease and to translate the current research into the therapeutic arena. In this review, we provide an overview of the current viral vector-based approaches and their potential. We discuss advances in platforms and injection routes that have been explored to enhance gene delivery toward kidney cells in animal models, and how these can fuel the development of viable gene therapy products for humans.

A noval noninvasive targeted therapy for osteosarcoma: the combination of LIFU and ultrasound-magnetic-mediated SPIO/TP53/PLGA nanobubble.

Osteosarcoma (OS) is the most common type of primary malignant bone tumor. Transducing a functional TP53 gene can effectively inhibit OS cell activity. Poly lactic acid-glycolic acid (PLGA) nanobubbles (NBs) mediated by focused ultrasound (US) can introduce exogenous genes into target cells in animal models, but this technique relies on the passive free diffusion of agents across the body. The inclusion of superparamagnetic iron oxide (SPIO) in microbubbles allows for magnetic-based tissue localization. A low-intensity-focused ultrasound (LIFU) instrument was developed at our institute, and different intensities of LIFU can either disrupt the NBs (RLI-LIFU) or exert cytocidal effects on the target tissues (RHI-LIFU). Based on these data, we performed US-magnetic-mediated TP53-NB destruction and investigated its ability to inhibit OS growth when combined with LIFU both in vitro and in vivo. Several SPIO/TP53/PLGA (STP) NB variants were prepared and characterized. For the in vitro experiments, HOS and MG63 cells were randomly assigned into five treatment groups. Cell proliferation and the expression of TP53 were detected by CCK8, qRT-PCR and Western blotting, respectively. In vivo, tumor-bearing nude mice were randomly assigned into seven treatment groups. The iron distribution of Perls' Prussian blue-stained tissue sections was determined by optical microscopy. TUNEL-DAPI was performed to examine apoptosis. TP53 expression was detected by qRT-PCR and immunohistochemistry. SPIO/TP53/PLGA NBs with a particle size of approximately 200nm were prepared successfully. For in vitro experiments, ultrasound-targeted transfection of TP53 overexpression in OS cells and efficient inhibition of OS proliferation have been demonstrated. Furthermore, in a tumor-bearing nude mouse model, RLI-LIFU-magnetic-mediated SPIO/TP53/PLGA NBs increased the transfection efficiency of the TP53 plasmid, resulting in apoptosis. Adding RHI-LIFU to the treatment regimen significantly increased the apoptosis of OS cells in vivo. Combining LIFU and US-magnetic-mediated SPIO/TP53/PLGA NB destruction is potentially a novel noninvasive and targeted therapy for OS.

Unlocking the potential of iPSC-derived immune cells: engineering iNK and iT cells for cutting-edge immunotherapy.

Induced pluripotent stem cells (iPSCs) have emerged as a revolutionary tool in cell therapies due to their ability to differentiate into various cell types, unlimited supply, and potential as off-the-shelf cell products. New advances in iPSC-derived immune cells have generated potent iNK and iT cells which showed robust killing of cancer cells in animal models and clinical trials. With the advent of advanced genome editing technologies that enable the development of highly engineered cells, here we outline 12 strategies to engineer iPSCs to overcome limitations and challenges of current cell-based immunotherapies, including safety switches, stealth edits, avoiding graft-versus-host disease (GvHD), targeting, reduced lymphodepletion, efficient differentiation, increased in vivo persistence, stemness, metabolic fitness, homing/trafficking, and overcoming suppressive tumor microenvironment and stromal cell barrier. With the development of advanced genome editing techniques, it is now possible to insert large DNA sequences into precise genomic locations without the need for DNA double strand breaks, enabling the potential for multiplexed knock out and insertion. These technological breakthroughs have made it possible to engineer complex cell therapy products at unprecedented speed and efficiency. The combination of iPSC derived iNK, iT and advanced gene editing techniques provides new opportunities and could lead to a new era for next generation of cell immunotherapies.

Characteristics of sintered calcium deficient hydroxyapatite scaffolds produced by 3D printing

This study investigates the parametric quality and reliability of 3D-printed scaffolds using a composite filament comprising a thermoplastic polymer and hydroxyapatite in 1:1 wt ratio. Employing a fused filament fabrication printer, we verified the 3D printing strategy, sintering temperature range, and preparation of hydroxyapatite scaffolds. To complete the study, in vitro cytotoxicity tests using animal and human cell models were conducted. The calcium-deficient hydroxyapatite (Ca/P ∼ 1.54) used in preparation of filament exhibited after sintering differences in crystalochemical phases in different rations, comparing to stoichiometric hydroxyapatite (Ca/P ≅ 1.67). The FFF process successfully produced scaffold macropores suitable for tissue vascularization (∼380 µm or smaller). Optimal sintering temperature for cell proliferation was identified at 1300 °C, especially effective for used investigated calcium-deficient hydroxyapatite. Cytotoxicity assessment with murine and human fibroblastic cells demonstrated differing behaviour, emphasizing the need for careful material property modifications for practical scaffold utilization.

Identification and Analysis of Myogenic Progenitors In Vivo During Acute Skeletal Muscle Injury by High-Dimensional Single-Cell Mass Cytometry.

Skeletal muscle regeneration is a dynamic process driven by adult muscle stem cells and their progeny. Mostly quiescent at a steady state, adult muscle stem cells become activated upon muscle injury. Following activation, they proliferate, and most of their progeny differentiate to generate fusion-competent muscle cells while the remaining self-renews to replenish the stem cell pool. While the identity of muscle stem cells was defined more than a decade ago, based on the co-expression of cell surface markers, myogenic progenitors were identified only recently using high-dimensional single-cell approaches. Here, we present a single-cell mass cytometry (cytometry by time of flight [CyTOF]) method to analyze stem cells and progenitor cells in acute muscle injury to resolve the cellular and molecular dynamics that unfold during muscle regeneration. This approach is based on the simultaneous detection of novel cell surface markers and key myogenic transcription factors whose dynamic expression enables the identification of activated stem cells and progenitor cell populations that represent landmarks of myogenesis. Importantly, a sorting strategy based on detecting cell surface markers CD9 and CD104 is described, enabling prospective isolation of muscle stem and progenitor cells using fluorescence-activated cell sorting (FACS) for in-depth studies of their function. Muscle progenitor cells provide a critical missing link to study the control of muscle stem cell fate, identify novel therapeutic targets for muscle diseases, and develop cell therapy applications for regenerative medicine. The approach presented here can be applied to study muscle stem and progenitor cells in vivo in response to perturbations, such as pharmacological interventions targeting specific signaling pathways. It can also be used to investigate the dynamics of muscle stem and progenitor cells in animal models of muscle diseases, advancing our understanding of stem cell diseases and accelerating the development of therapies.