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Prime editor, an editing tool based on the CRISPR/Cas9 system, allows for all 12 types of nucleotide exchanges and arbitrary indels in genomic sequences without the need for inducing DNA double-strand breaks. Despite its flexibility and precision, prime editing efficiency is still low and hindered by various factors such as target sites, editing types, and the length of the primer binding site. In this study, we developed a prime editing system by incorporating an RNA motif at the 3′ terminal of the pegRNA and integrating all twin prime editor factors into a single plasmid. These two strategies enhanced prime editing efficiency at target sites by up to 3.58-fold and 2.19-fold, respectively. Subsequently, enhanced prime editor was employed in goat cells and embryos to efficiently insert a 38 bp attB sequence into the Gt(ROSA)26Sor (Rosa26) and C-C motif chemokine receptor 5 (CCR5) loci. The enhanced prime editor can mediate 11.9% and 6.8% editing efficiency in parthenogenetic activation of embryos through embryo microinjection. In summary, our study introduces a modified prime editing system with improved editing and transfection efficiency, making it more suitable for inserting foreign sequences into primary cells and embryos. These results broaden the potential applications of prime editing technologies in the production of transgenic animals. [ABSTRACT FROM AUTHOR]

Simple Summary: Poor prognosis for advanced bladder cancer underlines the need for a better understanding of tumor resistance to treatment in relevant preclinical models. We developed new models of inducible and orthotopic muscle-invasive bladder cancer, with strong phenotypical and genetic resemblances to the human disease. These models enable real-time tracking of tumor progression and metastasis through bioluminescence imaging to facilitates the testing of new therapies. This valuable tool is important for advancing bladder cancer research and developing more effective treatments. Bladder cancer (BC) is the fourth most common cancer in men, with a poor patient prognosis for advanced disease. The poor survival of patients with muscle-invasive bladder cancer (MIBC) and metastatic status emphasizes the urgent need to develop new therapies. Lacking in the field of BC is the availability of relevant advanced BC mouse models, especially metastatic ones, that accurately recapitulate the complexities of human pathology to test and study new therapeutic strategies. Addressing this need, we developed a traceable mouse model of BC that expresses tumor-associated antigens within the context of advanced muscle-invasive BC. This novel system was achieved through the deletion of the tp53 and pten genes, alongside the incorporation of the fusion construct of Firefly luciferase (Luc) and the SIYRYYGL (SIY) T-cell antigen. We validate that the presence of the transgene did not impact on the development of the tumors while allowing us to measure tumor progression by bioluminescence. We show that the transgene did not influence the composition of the immune tumor microenvironment. More importantly, we report that this model was unresponsive to anti-PD-1 treatment, as in the majority of patients with BC. We also develop a new model based on the orthotopic injection of BC clonal cell lines derived from our first model. We demonstrate that this new model invades the muscle layer and has a metastasis development rate of 83%. The advantage of this model is that we can visualize tumor growth and metastasis development in vivo. These mouse models' characteristics, displaying many similarities with the human pathology, provide a valuable tool for tracking tumor progression, metastasis spread in vivo, and treatment resistance, as well as exploring fundamental and translational aspects of BC biology. This work contributes to the improvement in the landscape of mouse models of advanced BC for testing new therapeutic strategies. [ABSTRACT FROM AUTHOR]

For at least two reasons, the current transgenic animal models of Alzheimer's disease (AD) appear to be patently inadequate. They may be useful in many respects, the AD models; however, they are not. First, they are incapable of developing the full spectrum of the AD pathology. Second, they respond spectacularly well to drugs that are completely ineffective in the treatment of symptomatic AD. These observations indicate that both the transgenic animal models and the drugs faithfully reflect the theory that guided the design and development of both, the amyloid cascade hypothesis (ACH), and that both are inadequate because their underlying theory is. This conclusion necessitated the formulation of a new, all-encompassing theory of conventional AD—the ACH2.0. The two principal attributes of the ACH2.0 are the following. One, in conventional AD, the agent that causes the disease and drives its pathology is the intraneuronal amyloid-β (iAβ) produced in two distinctly different pathways. Two, following the commencement of AD, the bulk of Aβ is generated independently of Aβ protein precursor (AβPP) and is retained inside the neuron as iAβ. Within the framework of the ACH2.0, AβPP-derived iAβ accumulates physiologically in a lifelong process. It cannot reach levels required to support the progression of AD; it does, however, cause the disease. Indeed, conventional AD occurs if and when the levels of AβPP-derived iAβ cross the critical threshold, elicit the neuronal integrated stress response (ISR), and trigger the activation of the AβPP-independent iAβ generation pathway; the disease commences only when this pathway is operational. The iAβ produced in this pathway reaches levels sufficient to drive the AD pathology; it also propagates its own production and thus sustains the activity of the pathway and perpetuates its operation. The present study analyzes the reason underlying the evident inadequacy of the current transgenic animal models of AD. It concludes that they model, in fact, not Alzheimer's disease but rather the effects of the neuronal ISR sustained by AβPP-derived iAβ, that this is due to the lack of the operational AβPP-independent iAβ production pathway, and that this mechanism must be incorporated into any successful AD model faithfully emulating the disease. The study dissects the plausible molecular mechanisms of the AβPP-independent iAβ production and the pathways leading to their activation, and introduces the concept of conventional versus unconventional Alzheimer's disease. It also proposes the path forward, posits the principles of design of productive transgenic animal models of the disease, and describes the molecular details of their construction. [ABSTRACT FROM AUTHOR]

Simple Summary: In this study, we screened key signaling pathways regulating the formation of chicken PGCs using transcriptome sequencing and established a two-step induction model of PGCs in vitro to improve the optimization of their induction efficiency. Our results provide cellular materials for realizing the specific application of PGCs. In recent years, inducing pluripotent stem cells to differentiate into functional primordial germ cells (PGCs) in vitro has become an important method of obtaining a large number of PGCs. However, the instability and low induction efficiency of the in vitro PGC induction system restrict the application of PGCs in transgenic animal production, germplasm resource conservation and other fields. In this study, we successfully established a two-step induction model of chicken PGCs in vitro, which significantly improved the formation efficiency of PGC-like cells (PGCLCs). To further improve the PGC formation efficiency in vitro, 5025 differentially expressed genes (DEGs) were obtained between embryonic stem cells (ESCs) and PGCs through RNA-seq. GO and KEGG enrichment analysis revealed that signaling pathways such as BMP4, Wnt and Notch were significantly activated during PGC formation, similar to other species. In addition, we noted that cAMP was activated during PGC formation, while MAPK was suppressed. Based on the results of our analysis, we found that the PGC formation efficiency was significantly improved after activating Wnt and inhibiting MAPK, and was lower than after activating cAMP. To sum up, in this study, we successfully established a two-step induction model of chicken PGCs in vitro with high PGC formation efficiency, which lays a theoretical foundation for further demonstrating the regulatory mechanism of PGCs and realizing their specific applications. [ABSTRACT FROM AUTHOR]

A hallmark of Alzheimer's disease (AD) are the proteinaceous aggregates formed by the amyloid-beta peptide (Aβ) that is deposited inside the brain as amyloid plaques. The accumulation of aggregated Aβ may initiate or enhance pathologic processes in AD. According to the amyloid hypothesis, any agent that has the capability to inhibit Aβ aggregation and/or destroy amyloid plaques represents a potential disease-modifying drug. In 2023, a humanized IgG1 monoclonal antibody (lecanemab) against the Aβ-soluble protofibrils was approved by the US FDA for AD therapy, thus providing compelling support to the amyloid hypothesis. To acquire a deeper insight on the in vivo Aβ aggregation, various animal models, including aged herbivores and carnivores, non-human primates, transgenic rodents, fish and worms were widely exploited. This review is based on the recent data obtained using transgenic animal AD models and presents experimental verification of the critical role in Aβ aggregation seeding of the interactions between zinc ions, Aβ with the isomerized Asp7 (isoD7-Aβ) and the α4β2 nicotinic acetylcholine receptor. [ABSTRACT FROM AUTHOR]

Simple Summary: Gastric cancer (G.C.) is a lethal human malignancy, boasting the highest mortality rate among all cancers. Recent advances in its molecular pathology have unveiled a comprehensive genetic profile associated with the initiation and progression of G.C. These encompass the activation of KRAS, along with the inactivation or loss of E-cadherin, PTEN, and P53, forming the cornerstone for delving into the genetic and molecular underpinnings of this malignancy. We analyzed the effects of the loss of LKB1, an E-cadherin expression regulator, and PTEN, a negative regulator of the phosphoinositide 3-kinase (PI3K) signaling pathway, on G.C. development in mice. Our results indicated that these two genes act as suppressors in gastric carcinogenesis, and the loss of these genes promotes the development of G.C. in mice. The LKB1 and PTEN genes are critical in gastric cancer (G.C.) development. LKB1, a robust tumor suppressor gene, encodes a serine/threonine kinase that directly triggers the activation of AMPK—an integral cellular metabolic kinase. The role of the LKB1 pathway extends to maintaining the stability of epithelial junctions by regulating E-cadherin expression. Conversely, PTEN, a frequently mutated tumor suppressor gene in various human cancers, emerges as a pivotal negative regulator of the phosphoinositide 3-kinase (PI3K) signaling pathway. This study is set to leverage the H+/K+ ATPase Cre transgene strain to precisely target Cre recombinase expression at parietal cells within the stomach. This strategic maneuver seeks to selectively nullify the functions of both LKB1 and PTEN in a manner specific to the stomach, thereby instigating the development of G.C. in a fashion akin to human gastric adenocarcinoma. Moreover, this study endeavors to dissect the intricate ways in which these alterations contribute to the histopathologic advancement of gastric tumors, their potential for invasiveness and metastasis, their angiogenesis, and the evolving tumor stromal microenvironment. Our results show that conditional deletion of PTEN and LKB1 provides an ideal cancer microenvironment for G.C. tumorigenesis by promoting cancer cell proliferation, angiogenesis, and metastasis. [ABSTRACT FROM AUTHOR]

This work aims to clarify the effect of dietary polyunsaturated fatty acid (PUFA) intake on the adult brain affected by amyloid pathology. McGill-R-Thy1-APP transgenic (Tg) rat and 5xFAD Tg mouse models that represent earlier or later disease stages were employed. The animals were exposed to a control diet (CD) or an HFD based on corn oil, from young (rats) or adult (mice) ages for 24 or 10 weeks, respectively. In rats and mice, the HFD impaired reference memory in wild-type (WT) animals but did not worsen it in Tg, did not cause obesity, and did not increase triglycerides or glucose levels. Conversely, the HFD promoted stronger microglial activation in Tg vs. WT rats but had no effect on cerebral amyloid deposition. IFN-γ, IL-1β, and IL-6 plasma levels were increased in Tg rats, regardless of diet, while CXCL1 chemokine levels were increased in HFD-fed mice, regardless of genotype. Hippocampal 3-nitrotyrosine levels tended to increase in HFD-fed Tg rats but not in mice. Overall, an HFD with an elevated omega-6-to-omega-3 ratio as compared to the CD (25:1 vs. 8.4:1) did not aggravate the outcome of AD regardless of the stage of amyloid pathology, suggesting that many neurobiological processes relevant to AD are not directly dependent on PUFA intake. [ABSTRACT FROM AUTHOR]

Huntington's disease (HD) is an autosomal-dominant inherited neurological disorder caused by a genetic mutation in the IT15 gene. This neurodegenerative disorder is caused by a polyglutamine repeat expansion mutation in the widely expressed huntingtin (HTT) protein. HD is characterized by the degeneration of basal ganglia neurons and progressive cell death in intrinsic neurons of the striatum, accompanied by dementia and involuntary abnormal choreiform movements. Animal models have been extensively studied and have proven to be extremely valuable for therapeutic target evaluations. They reveal the hallmark of the age-dependent formation of aggregates or inclusions consisting of misfolded proteins. Animal models of HD have provided a therapeutic strategy to treat HD by suppressing mutant HTT (mHTT). Transgenic animal models have significantly increased our understanding of the molecular processes and pathophysiological mechanisms underlying the HD behavioral phenotype. Since effective therapies to cure or interrupt the course of the disease are not yet available, clinical research will have to make use of reliable animal models. This paper reviews the main studies of rodents as HD animal models, highlighting the neurological and behavioral differences between them. The choice of an animal model depends on the specific aspect of the disease to be investigated. Toxin-based models can still be useful, but most experimental hypotheses depend on success in a genetic model, whose choice is determined by the experimental question. There are many animal models showing similar HD symptoms or pathologies. They include chemical-induced HDs and genetic HDs, where cell-free and cell culture, lower organisms (such as yeast, Drosophila, C. elegans, zebrafish), rodents (mice, rats), and non-human primates are involved. These models provide accessible systems to study molecular pathogenesis and test potential treatments. For developing more effective pharmacological treatments, better animal models must be available and used to evaluate the efficacy of drugs. [ABSTRACT FROM AUTHOR]

Mitochondrial dysfunction contributes to numerous chronic diseases, and mitochondria are targets for various toxins and xenobiotics. Therefore, the development of drugs or therapeutic strategies targeting mitochondria is an important task in modern medicine. It is well known that the primary, although not the sole, function of mitochondria is ATP generation, which is achieved by coupled respiration. However, a high membrane potential can lead to uncontrolled reactive oxygen species (ROS) production and associated dysfunction. For over 50 years, scientists have been studying various synthetic uncouplers, and for more than 30 years, uncoupling proteins that are responsible for uncoupled respiration in mitochondria. Additionally, the proteins of the mitochondrial alternative respiratory pathway exist in plant mitochondria, allowing noncoupled respiration, in which electron flow is not associated with membrane potential formation. Over the past two decades, advances in genetic engineering have facilitated the creation of various cellular and animal models that simulate the effects of uncoupled and noncoupled respiration in different tissues under various disease conditions. In this review, we summarize and discuss the findings obtained from these transgenic models. We focus on the advantages and limitations of transgenic organisms, the observed physiological and biochemical changes, and the therapeutic potential of uncoupled and noncoupled respiration. [ABSTRACT FROM AUTHOR]

Endocannabinoids were implicated in a variety of pathological conditions including anxiety and are considered promising new targets for anxiolytic drug development. The optimism concerning the potentials of this system for anxiolysis is probably justified. However, the complexity of the mechanisms affected by endocannabinoids, and discrepant findings obtained with various experimental approaches makes the interpretation of research results difficult. Here, we review the anxiety-related effects of the three main interventions used to study the endocannabinoid system: pharmacological agents active at endocannabinoid-binding sites present on both the cell membrane and in the cytoplasm, genetic manipulations targeting cannabinoid receptors, and function-enhancers represented by inhibitors of endocannabinoid degradation and transport. Binding-site ligands provide inconsistent findings probably because they activate a multitude of mechanisms concomitantly. More robust findings were obtained with genetic manipulations and particularly with function enhancers, which heighten ongoing endocannabinoid activation rather than affecting all mechanisms indiscriminately. The enhancement of ongoing activity appears to ameliorate stress-induced anxiety without consistent effects on anxiety in general. Limited evidence suggests that this effect is achieved by promoting active coping styles in critical situations. These findings suggest that the functional enhancement of endocannabinoid signaling is a promising drug development target for stress-related anxiety disorders. [ABSTRACT FROM AUTHOR]

Antithrombin III is an important anticoagulant factor with anti-inflammatory properties. However, few studies have explored its anti-inflammatory actions in ATIII overexpressed transgenic animals. In this study, the dairy goats with mammary overexpression of ATIII were used to investigate their general health, milk quality and particularly their response to inflammatory challenge. The results showed that transgenic goats have a normal phenotype regarding their physiological and biochemical parameters, including whole blood cells, serum protein levels, total cholesterol, urea nitrogen, uric acid, and total bilirubin, compared to the WT. In addition, the quality of milk also improved in transgenic animals compared to the WT, as indicated by the increased milk fat and dry matter content and the reduced somatic cell numbers. Under the stimulation of an LPS injection, the transgenic goats had elevated contents of IGA, IGM and superoxide dismutase SOD, and had reduced proinflammatory cytokine release, including IL-6, TNF-α and IFN-β. A 16S rDNA sequencing analysis also showed that the transgenic animals had a similar compositions of gut microbiota to the WT goats under the stimulation of LPS injections. Mammary gland ATIII overexpression in dairy goats is a safe process, and it did not jeopardize the general health of the transgenic animals; moreover, the compositions of their gut microbiota also improved with the milk quality. The LPS stimulation study suggests that the increased ATIII expression may directly or indirectly suppress the inflammatory response to increase the resistance of transgenic animals to pathogen invasion. This will be explored in future studies. [ABSTRACT FROM AUTHOR]

The vascular endothelium is key target for immune and thrombotic responses that has to be controlled in successful xenotransplantation. Several genes were identified that, if induced or overexpressed, help to regulate the inflammatory response and preserve the transplanted organ function and metabolism. However, few studies addressed combined expression of such genes. The aim of this work was to evaluate in vivo the effects of the simultaneous expression of three human genes in a mouse generated using the multi-cistronic F2A technology. Male 3-month-old mice that express human heme oxygenase 1 (hHO-1), ecto-5′-nucleotidase (hE5NT), and ecto-nucleoside triphosphate diphosphohydrolase 1 (hENTPD1) (Transgenic) were compared to wild-type FVB mice (Control). Background analysis include extracellular nucleotide catabolism enzymes profile on the aortic surface, blood nucleotide concentration, and serum L-arginine metabolites. Furthermore, inflammatory stress induced by LPS in transgenic and control mice was used to characterize interleukin 6 (IL-6) and adhesion molecules endothelium permeability responses. Transgenic mice had significantly higher rates of extracellular adenosine triphosphate and adenosine monophosphate hydrolysis on the aortic surface in comparison to control. Increased levels of blood AMP and adenosine were also noticed in transgenics. Moreover, transgenic animals demonstrated the decrease in serum monomethyl-L-arginine level and a higher L-arginine/monomethyl-L-arginine ratio. Importantly, significantly decreased serum IL-6, and adhesion molecule levels were observed in transgenic mice in comparison to control after LPS treatment. Furthermore, reduced endothelial permeability in the LPS-treated transgenic mice was noted as compared to LPS-treated control. The human enzymes (hHO-1, hE5NT, hENTPD1) simultaneously encoded in transgenic mice demonstrated benefits in several biochemical and functional aspects of endothelium. This is consistent in use of this approach in the context of xenotransplantation. [ABSTRACT FROM AUTHOR]

One hundred and twenty-five years ago there was a lively discussion between Hungarian and Spanish neuroscientists on the nature of neural connections. The question was whether the neurofibrils run from one neuron to the next and connect neurons as a continuous network or the fibrils form an internal skeleton in the neurons and do not leave the cell; however, there is close contact between the neurons. About 50 years later, the invention of the electron microscope solved the problem. Close contacts between individual neurons were identified and named as synapses. In the following years, the need arose to explore distant connections between neuronal structures. Tracing techniques entered neuroscience. There are three major groups of tracers: (A) non-transsynaptic tracers used to find direct connections between two neuronal structures; (B) tracers passing gap junctions; (C) transsynaptic tracers passing synapses that are suitable to explore multineuronal circuits. According to the direction of the transport mechanism, the tracer may be ante- or retrograde. In this review, we focus on the ever-increasing number of fluorescent tracers that we have also used in our studies. The advantage of the use of these molecules is that the fluorescence of the tracer can be seen in histological sections without any other processes. Genes encoding fluorescent molecules can be inserted in various neuropeptide or neurotransmitter expressing transcriptomes. This makes it possible to study the anatomy, development or functional relations of these neuronal networks in transgenic animals. [ABSTRACT FROM AUTHOR]

Astaxanthin is a fascinating molecule with powerful antioxidant activity, synthesized exclusively by specific microorganisms and higher plants. To expand astaxanthin production, numerous studies have employed metabolic engineering to introduce and optimize astaxanthin biosynthetic pathways in microorganisms and plant hosts. Here, we report the metabolic engineering of animal cells in vitro to biosynthesize astaxanthin. This was accomplished through a two-step study to introduce the entire astaxanthin pathway into human embryonic kidney cells (HEK293T). First, we introduced the astaxanthin biosynthesis sub-pathway (Ast subp) using several genes encoding β-carotene ketolase and β-carotene hydroxylase enzymes to synthesize astaxanthin directly from β-carotene. Next, we introduced a β-carotene biosynthesis sub-pathway (β-Car subp) with selected genes involved in Ast subp to synthesize astaxanthin from geranylgeranyl diphosphate (GGPP). As a result, we unprecedentedly enabled HEK293T cells to biosynthesize free astaxanthin from GGPP with a concentration of 41.86 µg/g dry weight (DW), which represented 66.19% of the total ketocarotenoids (63.24 µg/g DW). Through optimization steps using critical factors in the astaxanthin biosynthetic process, a remarkable 4.14-fold increase in total ketocarotenoids (262.10 µg/g DW) was achieved, with astaxanthin constituting over 88.82%. This pioneering study holds significant implications for transgenic animals, potentially revolutionizing the global demand for astaxanthin, particularly within the aquaculture sector. [ABSTRACT FROM AUTHOR]

Simple Summary: Genetically modified farm animals have been actively used in a number of applications from testing gene functions to increasing production and economic value of livestock. To date, industrial genome engineering in production animals has been successfully applied to increase economically significant traits, reduce the health risk of animal products and express recombinant proteins for the needs of the pharmaceutical industry. Future applications of transgenic animals extend to producing xenografts for medical uses as well as many other promising areas. Here we review the perspectives of genome engineering in livestock from the technical point of view. Genome editing of farm animals has undeniable practical applications. It helps to improve production traits, enhances the economic value of livestock, and increases disease resistance. Gene-modified animals are also used for biomedical research and drug production and demonstrate the potential to be used as xenograft donors for humans. The recent discovery of site-specific nucleases that allow precision genome editing of a single-cell embryo (or embryonic stem cells) and the development of new embryological delivery manipulations have revolutionized the transgenesis field. These relatively new approaches have already proven to be efficient and reliable for genome engineering and have wide potential for use in agriculture. A number of advanced methodologies have been tested in laboratory models and might be considered for application in livestock animals. At the same time, these methods must meet the requirements of safety, efficiency and availability of their application for a wide range of farm animals. This review aims at covering a brief history of livestock animal genome engineering and outlines possible future directions to design optimal and cost-effective tools for transgenesis in farm species. [ABSTRACT FROM AUTHOR]

Simple Summary: The 5-year survival rate for women with metastatic breast cancer is 29%. Potential biomarker identification is important for new treatment modalities in affected patients. Previous studies have shown that Parathyroid hormone-related peptide (PTHrP) plays a critical role in breast cancer growth and metastasis. Our study aimed to use a genetically modified breast cancer mouse model to precisely examine the role of PTHrP from early primary breast cancer initiation to late progression. We identified a novel long non-coding RNA (lncRNA), a new target for fatty acid metabolism that can be regulated via PTHrP in our unique mouse breast cancer model. We confirmed that a potential human lncRNA, OLMALINC, plays a similar role in fatty acid metabolism that can be regulated via PTHrP and validated our mouse findings in human breast cancer cell lines. Genetically engineered mouse models provide valuable tools to study the molecular metabolism for breast cancer progression. Parathyroid hormone-related peptide (PTHrP) is the primary cause of malignancy-associated hypercalcemia (MAH). We previously showed that PTHrP ablation, in the MMTV-PyMT murine model of breast cancer (BC) progression, can dramatically prolong tumor latency, slow tumor growth, and prevent metastatic spread. However, the signaling mechanisms using lineage tracing have not yet been carefully analyzed. Here, we generated Pthrpflox/flox; Cre+ mT/mG mice (KO) and Pthrpwt/wt; Cre+ mT/mG tumor mice (WT) to examine the signaling pathways under the control of PTHrP from the early to late stages of tumorigenesis. GFP+ mammary epithelial cells were further enriched for subsequent RNA sequencing (RNAseq) analyses. We observed significant upregulation of cell cycle signaling and fatty acid metabolism in PTHrP WT tumors, which are linked to tumor initiation and progression. Next, we observed that the expression levels of a novel lncRNA, GM50337, along with stearoyl-Coenzyme A desaturase 1 (Scd1) are significantly upregulated in PTHrP WT but not in KO tumors. We further validated a potential human orthologue lncRNA, OLMALINC, together with SCD1 that can be regulated via PTHrP in human BC cell lines. In conclusion, these novel findings could be used to develop targeted strategies for the treatment of BC and its metastatic complications. [ABSTRACT FROM AUTHOR]

Gram-negative bacterial infections pose a significant threat to public health. Toll-like receptor 4 (TLR4) recognizes bacterial lipopolysaccharide (LPS) and induces innate immune responses, autophagy, and cell death, which have major impacts on the body's physiological homeostasis. However, the role of TLR4 in bacterial LPS-induced autophagy and apoptosis in large mammals, which are closer to humans than rodents in many physiological characteristics, remains unknown. So far, few reports focus on the relationship between TLR, autophagy, and apoptosis in large mammal levels, and we urgently need more tools to further explore their crosstalk. Here, we generated a TLR4-enriched mammal model (sheep) and found that a high-dose LPS treatment blocked autophagic degradation and caused strong innate immune responses and severe apoptosis in monocytes/macrophages of transgenic offspring. Excessive accumulation of autophagosomes/autolysosomes might contribute to LPS-induced apoptosis in monocytes/macrophages of transgenic animals. Further study demonstrated that inhibiting TLR4 downstream NF-κB or p38 MAPK signaling pathways reversed the LPS-induced autophagy activity and apoptosis. These results indicate that the elevated TLR4 aggravates LPS-induced monocytes/macrophages apoptosis by leading to lysosomal dysfunction and impaired autophagic flux, which is associated with TLR4 downstream NF-κB and MAPK signaling pathways. This study provides a novel TLR4-enriched mammal model to study its potential effects on autophagy activity, inflammation, oxidative stress, and cell death. These findings also enrich the biological functions of TLR4 and provide powerful evidence for bacterial infection. [ABSTRACT FROM AUTHOR]

Research into genetic and physiological mechanisms of widespread disorders such as arterial hypertension as well as neuropsychiatric and other human diseases is urgently needed in academic and practical medicine and in the field of biology. Nevertheless, such studies have many limitations and pose difficulties that can be overcome by using animal models. To date, for the purposes of creating animal models of human pathologies, several approaches have been used: pharmacological/chemical intervention; surgical procedures; genetic technologies for creating transgenic animals, knockouts, or knockdowns; and breeding. Although some of these approaches are good for certain research aims, they have many drawbacks, the greatest being a strong perturbation (in a biological system) that, along with the expected effect, exerts side effects in the study. Therefore, for investigating the pathogenesis of a disease, models obtained using genetic selection for a target trait are of high value as this approach allows for the creation of a model with a "natural" manifestation of the pathology. In this review, three rat models are described: ISIAH rats (arterial hypertension), GC rats (catatonia), and PM rats (audiogenic epilepsy), which are developed by breeding in the Laboratory of Evolutionary Genetics at the Institute of Cytology and Genetics (the Siberian Branch of the Russian Academy of Sciences). [ABSTRACT FROM AUTHOR]

Simple Summary: ATRX mutations occur in up to 17% of human pancreatic neuroendocrine tumours (PanNETs), and recent evidence points towards its inability to drive PanNET formation in mouse pancreas while predisposing individuals to inflammageing. Aiming to explore the additional non-tumourigenic consequences of Atrx deletion, we characterised an aged series of Atrx conditional disruption in β cells using the Pdx1 promoter. Homozygous mice (P.AtrxHOM) exhibited obesity, diabetes, glucose intolerance, and pancreatic adiposity at a higher extent than age- and sex-matched controls (P.AtrxWT). Atrx loss was recently ascertained as insufficient to drive pancreatic neuroendocrine tumour (PanNET) formation in mice islets. We have identified a preponderant role of Atrx in the endocrine dysfunction in a Rip-Cre;AtrxKO genetically engineered mouse model (GEMM). To validate the impact of a different Cre-driver line, we used similar methodologies and characterised the Pdx1-Cre;AtrxKO (P.AtrxKO) GEMM to search for PanNET formation and endocrine fitness disruption for a period of up to 24 months. Male and female mice presented different phenotypes. Compared to P.AtrxWT, P.AtrxHOM males were heavier during the entire study period, hyperglycaemic between 3 and 12 mo., and glucose intolerant only from 6 mo.; in contrast, P.AtrxHOM females started exhibiting increased weight gains later (after 6 mo.), but diabetes or glucose intolerance was detected by 3 mo. Overall, all studied mice were overweight or obese from early ages, which challenged the histopathological evaluation of the pancreas and liver, especially after 12 mo. Noteworthily, losing Atrx predisposed mice to an increase in intrapancreatic fatty infiltration (FI), peripancreatic fat deposition, and macrovesicular steatosis. As expected, no animal developed PanNETs. An obese diabetic GEMM of disrupted Atrx is presented as potentially useful for metabolic studies and as a putative candidate for inserting additional tumourigenic genetic events. [ABSTRACT FROM AUTHOR]

Simple Summary: The current article provides an overview of the significance of using various mouse and zebrafish models in cancer research. In addition, this review discusses an interdisciplinary 'Team Medicine' approach that has aided in enhancing our understanding of carcinogenesis and establishing new therapeutic approaches. The biological course of human malignancies, preclinical research on prospective cancer treatments, and cancer prevention benefit significantly from experimental animal models. The molecular mechanisms underlying tumor growth, progression, metastasis, maintenance, and acquisition of chemo-resistance have received significant attention in the study using different types of mice cancer models. Zebrafish have been recommended as a potential model to investigate human cancer because of their suitability for in vivo imaging, fast development, chemical screening, and adaptable genetics. The zebrafish's forward genetics and vertebrate biology make it a model system with immense potential for understanding carcinogenesis. The significance of applying various animal models in studying cancer development and progression has been well proven. The current review provides a comprehensive description of how these diverse models may be applied productively based on the scientific challenges that need to be addressed. Animal models have been utilized for decades to investigate the causes of human diseases and provide platforms for testing novel therapies. Indeed, breakthrough advances in genetically engineered mouse (GEM) models and xenograft transplantation technologies have dramatically benefited in elucidating the mechanisms underlying the pathogenesis of multiple diseases, including cancer. The currently available GEM models have been employed to assess specific genetic changes that underlay many features of carcinogenesis, including variations in tumor cell proliferation, apoptosis, invasion, metastasis, angiogenesis, and drug resistance. In addition, mice models render it easier to locate tumor biomarkers for the recognition, prognosis, and surveillance of cancer progression and recurrence. Furthermore, the patient-derived xenograft (PDX) model, which involves the direct surgical transfer of fresh human tumor samples to immunodeficient mice, has contributed significantly to advancing the field of drug discovery and therapeutics. Here, we provide a synopsis of mouse and zebrafish models used in cancer research as well as an interdisciplinary 'Team Medicine' approach that has not only accelerated our understanding of varied aspects of carcinogenesis but has also been instrumental in developing novel therapeutic strategies. [ABSTRACT FROM AUTHOR]

Simple Summary: Adrenocortical cancer is a very rare form of endocrine cancer with dismal prognosis. Preclinical models such as cell lines, organoids, and mouse models are essential for both improving basic understanding of this disease and developing treatments. Herein, we review a currently available model for adrenocortical cancers, with a special focus on adrenocortical carcinoma. Recent developments in in vitro models have included cell and 3D culture models with improved recapitulation of the tumor microenvironment and genetics. We hope to improve visibility and access to these models through this review. Adrenocortical cancer is an aggressive endocrine malignancy with an incidence of 0.72 to 1.02 per million people/year, and a very poor prognosis with a five-year survival rate of 22%. As an orphan disease, clinical data are scarce, meaning that drug development and mechanistic research depend especially on preclinical models. While a single human ACC cell line was available for the last three decades, over the last five years, many new in vitro and in vivo preclinical models have been generated. Herein, we review both in vitro (cell lines, spheroids, and organoids) and in vivo (xenograft and genetically engineered mouse) models. Striking leaps have been made in terms of the preclinical models of ACC, and there are now several modern models available publicly and in repositories for research in this area. [ABSTRACT FROM AUTHOR]

Simple Summary: Different reproductive biotechnologies have been applied to sheep, such as cloning, which has been successfully applied in this species. In this context, the aim of the present study was to carry out a bibliometric analysis of the scientific literature on cloning applied to sheep reproduction, since the first report was published, to identify the most cited articles, main authors and collaboration among them, published journals, institutions with more published papers, most prolific countries and the network collaboration among them, and research topics. This study collected bibliographic data from 124 papers relating to cloning of sheep. The articles that were cited more often addressed topics related to the generation of transgenic animals, recovery of wild species, and xenotransplants. So far, no bibliometric studies have been conducted about cloning of sheep. Somatic cell nuclear transfer (SCNT) is a reproductive biotechnology with great potential in the reproduction of different species of zootechnical interest, including sheep. This study aimed to carry out a bibliometric analysis of scientific papers published on the application of SCNT in sheep reproduction during the period 1997–2023. The search involved the Science Citation Index Expanded and Social Sciences Citation Index databases of the main collection of the Web of Sciences with different descriptors. A total of 124 scientific papers were analyzed for different bibliometric indicators using the VOSviewer software. Since 2001, the number of SCNT-related papers that have been published concerning sheep reproduction has increased and it has fluctuated in ensuing years. The main authors, research groups, institutions, countries, papers, and journals with the highest number of papers related to the application of SCNT in sheep reproduction were identified, as well as the topics that address the research papers according to the terms: somatic cell, embryo, oocyte, gene expression, SCNT, and sheep. [ABSTRACT FROM AUTHOR]

The P2X7 receptor is a trimeric ligand-gated cation channel activated by extracellular adenosine 5′-triphosphate. The study of animals has greatly advanced the investigation of P2X7 and helped to establish the numerous physiological and pathophysiological roles of this receptor in human health and disease. Following a short overview of the P2X7 distribution, roles and functional properties, this article discusses how animal models have contributed to the generation of P2X7-specific antibodies and nanobodies (including biologics), recombinant receptors and radioligands to study P2X7 as well as to the pharmacokinetic testing of P2X7 antagonists. This article then outlines how mouse and rat models have been used to study P2X7. These sections include discussions on preclinical disease models, polymorphic P2X7 variants, P2X7 knockout mice (including bone marrow chimeras and conditional knockouts), P2X7 reporter mice, humanized P2X7 mice and P2X7 knockout rats. Finally, this article reviews the limited number of studies involving guinea pigs, rabbits, monkeys (rhesus macaques), dogs, cats, zebrafish, and other fish species (seabream, ayu sweetfish, rainbow trout and Japanese flounder) to study P2X7. [ABSTRACT FROM AUTHOR]

A great deal of evidence supports the inevitable importance of spinal glycinergic inhibition in the development of chronic pain conditions. However, it remains unclear how glycinergic neurons contribute to the formation of spinal neural circuits underlying pain-related information processing. Thus, we intended to explore the synaptic targets of spinal glycinergic neurons in the pain processing region (laminae I–III) of the spinal dorsal horn by combining transgenic technology with immunocytochemistry and in situ hybridization accompanied by light and electron microscopy. First, our results suggest that, in addition to neurons in laminae I–III, glycinergic neurons with cell bodies in lamina IV may contribute substantially to spinal pain processing. On the one hand, we show that glycine transporter 2 immunostained glycinergic axon terminals target almost all types of excitatory and inhibitory interneurons identified by their neuronal markers in laminae I–III. Thus, glycinergic postsynaptic inhibition, including glycinergic inhibition of inhibitory interneurons, must be a common functional mechanism of spinal pain processing. On the other hand, our results demonstrate that glycine transporter 2 containing axon terminals target only specific subsets of axon terminals in laminae I–III, including nonpeptidergic nociceptive C fibers binding IB4 and nonnociceptive myelinated A fibers immunoreactive for type 1 vesicular glutamate transporter, indicating that glycinergic presynaptic inhibition may be important for targeting functionally specific subpopulations of primary afferent inputs. [ABSTRACT FROM AUTHOR]

A T cell receptor (TCR) consists of α- and β-chains. Accumulating evidence suggests that some TCRs possess chain centricity, i.e., either of the hemi-chains can dominate in antigen recognition and dictate the TCR's specificity. The introduction of TCRα/β into naive lymphocytes generates antigen-specific T cells that are ready to perform their functions. Transgenesis of the dominant active TCRα creates transgenic animals with improved anti-tumor immune control, and adoptive immunotherapy with TCRα-transduced T cells provides resistance to infections. However, the potential detrimental effects of the dominant hemi-chain TCR's expression in transgenic animals have not been well investigated. Here, we analyzed, in detail, the functional status of the immune system of recently generated 1D1a transgenic mice expressing the dominant active TCRα specific to the H2-Kb molecule. In their age dynamics, neither autoimmunity due to the random pairing of transgenic TCRα with endogenous TCRβ variants nor significant disturbances in systemic homeostasis were detected in these mice. Although the specific immune response was considerably enhanced in 1D1a mice, responses to third-party alloantigens were not compromised, indicating that the expression of dominant active TCRα did not limit immune reactivity in transgenic mice. Our data suggest that TCRα transgene expression could delay thymic involution and maintain TCRβ repertoire diversity in old transgenic mice. The detected changes in the systemic homeostasis in 1D1a transgenic mice, which are minor and primarily transient, may indicate variations in the ontogeny of wild-type and transgenic mouse lines. [ABSTRACT FROM AUTHOR]

Poultry are one of the most valuable resources for human society. They are also recognized as a powerful experimental animal for basic research on embryogenesis. Demands for the supply of low-allergen eggs and bioreactors have increased with the development of programmable genome editing technology. The CRISPR/Cas9 system has recently been used to produce transgenic animals and various animals in the agricultural industry and has also been successfully adopted for the modification of chicken and quail genomes. In this review, we describe the successful establishment of genome-edited lines combined with germline chimera production systems mediated by primordial germ cells and by viral infection in poultry. The avian intracytoplasmic sperm injection (ICSI) system that we previously established and recent advances in ICSI for genome editing are also summarized. [ABSTRACT FROM AUTHOR]

Simple Summary: Synovial sarcoma (SyS) is a rare malignant tumor mainly occurring in children, adolescents, and young adults. SyS displays the pathognomonic t(X;18) translocation resulting in the SS18-SSX fusion protein being able to interact with both the BAF enhancer complexes and polycomb repressor complexes, and either activate or repress gene transcription, resulting in genome-wide epigenetic deregulation and altered gene expression. This review analyzes the different experimental in vivo models for SyS research: (I) conditional transgenic mouse models expressing the SS18-SSX fusion protein that, alone or combined with some of the few other recurrent alterations (gains in BCL2, Wnt-β-catenin signaling, FGFR family, or loss of PTEN and SMARCB1), spontaneously develop SyS; (II) SyS patient-derived xenografts (PDX) established in immunodeficient mice; (III) SyS cell lines and cell line-derived xenografts. SyS preclinical models are greatly contributing to the disclosure of additional vulnerabilities and to the development of new therapeutic approaches for SyS. Synovial sarcomas (SyS) are rare malignant tumors predominantly affecting children, adolescents, and young adults. The genetic hallmark of SyS is the t(X;18) translocation encoding the SS18-SSX fusion gene. The fusion protein interacts with both the BAF enhancer and polycomb repressor complexes, and either activates or represses target gene transcription, resulting in genome-wide epigenetic perturbations and altered gene expression. Several experimental in in vivo models, including conditional transgenic mouse models expressing the SS18-SSX fusion protein and spontaneously developing SyS, are available. In addition, patient-derived xenografts have been estab-lished in immunodeficient mice, faithfully reproducing the complex clinical heterogeneity. This review focuses on the main molecular features of SyS and the related preclinical in vivo and in vitro models. We will analyze the different conditional SyS mouse models that, after combination with some of the few other recurrent alterations, such as gains in BCL2, Wnt-β-catenin signaling, FGFR family, or loss of PTEN and SMARCB1, have provided additional insight into the mechanisms of synovial sarcomagenesis. The recent advancements in the understanding of SyS biology and improvements in preclinical modeling pave the way to the development of new epigenetic drugs and immunotherapeutic approaches conducive to new treatment options. [ABSTRACT FROM AUTHOR]

Simple Summary: The incidence of prostate cancer is increasing because of the aging population. Evidence suggests that diets rich in bioactive polyphenols can reduce the incidence of prostate cancer. The aim of this work was to investigate the potential of gnetin C, a compound found in the melinjo plant and commonly used in Indonesian foods, to block prostate cancer progression. To this end, we evaluated the anticancer efficacy of gnetin C-supplemented diets in a unique and adequate high-risk premalignant prostate cancer transgenic mouse model. Our results indicate that a gnetin C-supplemented diet reduces the progression of prostate cancer by reducing the proliferation of cells, inflammation, and the formation of blood vessels. The finding that a gnetin C-supplemented diet effectively blocks tumor progression in a preclinical mouse model may be exploited to initiate chemoprevention trials for novel nutritional interception for untreated patients under active surveillance. Nutritional chemoprevention is particularly suitable for prostate cancer. Gnetin C, a resveratrol dimer found abundantly in the melinjo plant (Gnetum gnemon), may possess more potent biological properties compared to other stilbenes. We examined the effects of gnetin C in a high-risk premalignant transgenic mouse model overexpressing tumor-promoting metastasis-associated protein 1 (MTA1) on the background of Pten heterozygosity (R26MTA1; Pten+/f; Pb-Cre+). Mice were fed diets supplemented with the following compounds: pterostilbene (70 mg/kg diet); gnetin C, high dose (70 mg/kg diet); and gnetin C, low dose (35 mg/kg diet). Prostate tissues were isolated after 17 weeks and examined for histopathology and molecular markers. Serum was analyzed for cytokine expression. Gnetin C-supplemented diets substantially delayed the progression of preneoplastic lesions compared to other groups. Prostate tissues from gnetin C-fed mice showed favorable histopathology, with decreased severity and number of prostatic intraepithelial neoplasia (PIN) foci, reduced proliferation, and angiogenesis. A decreased level of MTA1, concurrent with the trend of increasing phosphatase and tensin homolog expression and reduced interleukin 2 (IL-2) levels in sera, were also detected in gnetin C-fed mice. Importantly, gnetin C did not exert any visible toxicity in mice. Our findings demonstrate that a gnetin C-supplemented diet effectively blocks MTA1-promoted tumor progression activity in high-risk premalignant prostate cancer, which indicates its potential as a novel form of nutritional interception for prostate cancer chemoprevention. [ABSTRACT FROM AUTHOR]

Simple Summary: Neuroendocrine neoplasia comprise many distinct and rare subtypes of cancers. Preclinical models are essential for improving understanding of these diseases because clinical data is scarce. We review available preclinical models across a wide spectrum of neuroendocrine neoplasia (including those affecting the lungs, gastrointestinal system, prostate, and adrenal glands). We consider models of varying complexity and accuracy, covering both in vitro models such as cell lines and 3D models, and in vivo models such as xenografts and genetically-engineered mouse models. Better access and understanding of these models as provided in this work will help to enable research into pathology and treatment across the spectrum of neuroendocrine neoplasia. Neuroendocrine neoplasia (NENs) are a complex and heterogeneous group of cancers that can arise from neuroendocrine tissues throughout the body and differentiate them from other tumors. Their low incidence and high diversity make many of them orphan conditions characterized by a low incidence and few dedicated clinical trials. Study of the molecular and genetic nature of these diseases is limited in comparison to more common cancers and more dependent on preclinical models, including both in vitro models (such as cell lines and 3D models) and in vivo models (such as patient derived xenografts (PDXs) and genetically-engineered mouse models (GEMMs)). While preclinical models do not fully recapitulate the nature of these cancers in patients, they are useful tools in investigation of the basic biology and early-stage investigation for evaluation of treatments for these cancers. We review available preclinical models for each type of NEN and discuss their history as well as their current use and translation. [ABSTRACT FROM AUTHOR]

The biochemical mechanisms of cell injury and myocardial cell death after myocardial infarction remain unresolved. Cyclooxygenase 2 (COX-2), a key enzyme in prostanoid synthesis, is expressed in human ischemic myocardium and dilated cardiomyopathy, but it is absent in healthy hearts. To assess the role of COX-2 in cardiovascular physiopathology, we developed transgenic mice that constitutively express functional human COX-2 in cardiomyocytes under the control of the α-myosin heavy chain promoter. These animals had no apparent phenotype but were protected against ischemia-reperfusion injury in isolated hearts, with enhanced functional recovery and diminished cellular necrosis. To further explore the phenotype of this animal model, we carried out a differential proteome analysis of wild-type vs. transgenic cardiomyocytes. The results revealed a tissue-specific proteomic profile dominated by mitochondrial proteins. In particular, an increased expression of respiratory chain complex IV proteins was observed. This correlated with increased catalytic activity, enhanced respiratory capacity, and increased ATP levels in the heart of COX-2 transgenic mice. These data suggest a new link between COX-2 and mitochondria, which might contribute to the protective cardiac effects of COX-2 against ischemia-reperfusion injury. [ABSTRACT FROM AUTHOR]

Simple Summary: The stage of a tumor during cancer intervention is the most crucial factor that determines the treatment regimen. Several bioactive natural compounds have shown potential to inhibit prostate cancer growth and progression; however, there is a dearth of studies that explore their efficacy at different stages of tumorigenesis. This knowledge gap prevents researchers from fully exploiting the anti-cancer potential of these beneficial compounds. Accordingly, our present study focused on explicating the 'stage-specific' efficacy of the bioactive food component 'inositol hexaphosphate (IP6, phytic acid)' against PCa initiation, growth, and progression in the transgenic adenocarcinoma of the mouse prostate TRAMP model. Results indicated that IP6 feeding during initial stages of cancer development prevents progression of prostatic intraepithelial neoplasia lesions to adenocarcinoma, and IP6 feeding during late stage of the disease reduces tumor growth and prevents its progression to advanced stage of the disease. Thus, IP6 intervention is beneficial during all stages of prostate tumorigenesis. Herein, we assessed the stage-specific efficacy of inositol hexaphosphate (IP6, phytic acid), a bioactive food component, on prostate cancer (PCa) growth and progression in a transgenic mouse model of prostate cancer (TRAMP). Starting at 4, 12, 20, and 30 weeks of age, male TRAMP mice were fed either regular drinking water or 2% IP6 in water for ~8–15 weeks. Pathological assessments at study endpoint indicated that tumor grade is arrested at earlier stages by IP6 treatment; IP6 also prevented progression to more advanced forms of the disease (~55–70% decrease in moderately and poorly differentiated adenocarcinoma incidence was observed in advanced stage TRAMP cohorts). Next, we determined whether the protective effects of IP6 are mediated via its effect on the expansion of the cancer stem cells (CSCs) pool; results indicated that the anti-PCa effects of IP6 are associated with its potential to eradicate the PCa CSC pool in TRAMP prostate tumors. Furthermore, in vitro assays corroborated the above findings as IP6 decreased the % of floating PC-3 prostaspheres (self-renewal of CSCs) by ~90%. Together, these findings suggest the multifaceted chemopreventive-translational potential of IP6 intervention in suppressing the growth and progression of PCa and controlling this malignancy at an early stage. [ABSTRACT FROM AUTHOR]

Myocilin is an enigmatic glaucoma-associated glycoprotein whose biological role remains incompletely understood. To gain novel insight into its normal function, we used transposon-mediated transgenesis to generate the first zebrafish line stably overexpressing myocilin [Tg(actb1:myoc-2A-mCherry)]. qPCR showed an approximately four-fold increased myocilin expression in transgenic zebrafish embryos (144 hpf). Adult (13 months old) transgenic animals displayed variable and age-dependent ocular anterior segment alterations. Almost 60% of two-year-old male, but not female, transgenic zebrafish developed enlarged eyes with severe asymmetrical and variable abnormalities in the anterior segment, characterized by corneal limbus hypertrophy, and thickening of the cornea, iris, annular ligament and lens capsule. The most severe phenotype presented small or absent ocular anterior chamber and pupils, due to iris overgrowth along with dysplastic retinal growth and optic nerve hypertrophy. Immunohistochemistry revealed increased presence of myocilin in most altered ocular tissues of adult transgenic animals, as well as signs of retinal gliosis and expanded ganglion cells and nerve fibers. The preliminary results indicate that these cells contributed to retinal dysplasia. Visual impairment was demonstrated in all old male transgenic zebrafish. Transcriptomic analysis of the abnormal transgenic eyes identified disrupted expression of genes involved in lens, muscular and extracellular matrix activities, among other processes. In summary, the developed transgenic zebrafish provides a new tool to investigate this puzzling protein and provides evidence for the role of zebrafish myocilin in ocular anterior segment and retinal biology, through the influence of extracellular matrix organization and cellular proliferation. [ABSTRACT FROM AUTHOR]

(1) The neurotrophic protein S100B is a marker of brain injury and has been associated with neuroregeneration. In S100Btg mice rendering 12 copies of the murine S100B gene we evaluated whether S100B may serve as a treatment option. (2) In juvenile, adult, and one-year-old S100Btg mice (female and male; n = 8 per group), progenitor cell proliferation was quantified in the subgranular zone (SGZ) and the granular cell layer (GCL) of the dentate gyrus with the proliferative marker Ki67 and BrdU (50 mg/kg). Concomitant signaling was quantified utilizing glial fibrillary acidic protein (GFAP), apolipoprotein E (ApoE), brain-derived neurotrophic factor (BDNF), and the receptor for advanced glycation end products (RAGE) immunohistochemistry. (3) Progenitor cell proliferation in the SGZ and migration to the GCL was enhanced. Hippocampal GFAP was reduced in one-year-old S100Btg mice. ApoE in the hippocampus and frontal cortex of male and BDNF in the frontal cortex of female S100Btg mice was reduced. RAGE was not affected. (4) Enhanced hippocampal neurogenesis in S100Btg mice was not accompanied by reactive astrogliosis. Sex- and brain region-specific variations of ApoE and BDNF require further elucidations. Our data reinforce the importance of this S100Btg model in evaluating the role of S100B in neuroregenerative medicine. [ABSTRACT FROM AUTHOR]

Simple Summary: Intraductal papillary mucinous neoplasms (IPMN) are common premalignant precursors to pancreatic ductal adenocarcinoma (PDAC). However, current guidelines for clinical management of IPMN are suboptimal and limited to information obtained through anatomical imaging and cytology of biopsy samples. Our results have suggested that diffusion-weighted MRI, an imaging method sensitive to tumor cell density, architecture, and microenvironment is able to differentiate mouse models of IPMN versus PDAC. KPC (KrasG12D:Trp53R172H:Pdx1-Cre) and CKS (KrasG12D:Smad4L/L:Ptf1a-Cre) mice are genetically engineered mouse (GEM) models that capture features of human pancreatic ductal adenocarcinoma (PDAC) and intraductal papillary mucinous neoplasms (IPMN), respectively. We compared these autochthonous tumors using quantitative imaging metrics from diffusion-weighted MRI (DW-MRI) and dynamic contrast enhanced (DCE)-MRI in reference to quantitative histological metrics including cell density, fibrosis, and microvasculature density. Our results revealed distinct DW-MRI metrics between the KPC vs. CKS model (mimicking human PDAC vs. IPMN lesion): the apparent diffusion coefficient (ADC) of CKS tumors is significantly higher than that of KPC, with little overlap (mean ± SD 2.24 ± 0.2 vs. 1.66 ± 0.2 , p < 10 − 10 ) despite intratumor and intertumor variability. Kurtosis index (KI) is also distinctively separated in the two models. DW imaging metrics are consistent with growth pattern, cell density, and the cystic nature of the CKS tumors. Coregistration of ex vivo ADC maps with H&E-stained sections allowed for regional comparison and showed a correlation between local cell density and ADC value. In conclusion, studies in GEM models demonstrate the potential utility of diffusion-weighted MRI metrics for distinguishing pancreatic cancer from benign pancreatic cysts such as IPMN. [ABSTRACT FROM AUTHOR]

Simple Summary: Lung cancer that is driven by mutations in the epidermal growth factor receptor (EGFR) is currently treated with tyrosine kinase inhibitors (TKIs). Although patients initially respond well to TKI treatment, drug resistance against EGFR-targeted therapy emerges. Attempts to combine immunotherapy with EGFR-targeted treatment to prolong response rates or prevent the development of resistances have been limited due to insufficient knowledge about the effects of targeted therapy on the tumour microenvironment (TME) in EGFR-driven tumours and tumour-infiltrating immune cells. The aims of this study were to improve our understanding on the impact of EGFR inhibition on the immune response in EGFR-driven lung cancer and, furthermore, to gain insights into the impact of combining targeted therapy with immunotherapy on the TME. EGFR-driven non-small-cell lung cancer (NSCLC) patients are currently treated with TKIs targeting EGFR, such as erlotinib or osimertinib. Despite a promising initial response to TKI treatment, most patients gain resistance to oncogene-targeted therapy, and tumours progress. With the development of inhibitors against immune checkpoints, such as PD-1, that mediate an immunosuppressive microenvironment, immunotherapy approaches attempt to restore a proinflammatory immune response in tumours. However, this strategy has shown only limited benefits in EGFR-driven NSCLC. Approaches combining EGFR inhibition with immunotherapy to stimulate the immune response and overcome resistance to therapy have been limited due to insufficient understanding about the effect of EGFR-targeting treatment on the immune cells in the TME. Here, we investigate the impact of EGFR inhibition by erlotinib on the TME and its effect on the antitumour response of the immune cell infiltrate. For this purpose, we used a transgenic conditional mouse model to study the immunological profile in EGFR-driven NSCLC tumours. We found that EGFR inhibition mediated a higher infiltration of immune cells and increased local proliferation of T-cells in the tumours. Moreover, inhibiting EGFR signalling led to increased activation of immune cells in the TME. Most strikingly, combined simultaneous blockade of EGFR and anti-PD-1 (aPD-1) enhanced tumour treatment response in a transgenic mouse model of EGFR-driven NSCLC. Thus, our findings show that EGFR inhibition promotes an active and proinflammatory immune cell infiltrate in the TME while improving response to immune checkpoint inhibitors in EGFR-driven NSCLC. [ABSTRACT FROM AUTHOR]

Simple Summary: Many patients with chronic lymphocytic leukemia (CLL) still fail current therapies. CD73 is a novel therapeutic target for solid tumors, but its role in CLL remains unclear. The aim of our study was to investigate the therapeutic potential of targeting CD73 in CLL. Using genetically engineered mice, our study reports a pro-leukemic role for CD73 in an autochthonous mouse model of CLL. Furthermore, we observed an association between PD-L1 expression on CLL cells and adenosine signaling according to sex. Our findings provide a rationale for targeting CD73 in CLL in combination with anti-PD-1/PD-L1 immunotherapies and suggest that sex may contribute to responses to adenosine-targeting agents. The ecto-nucleotidase CD73 is an important immune checkpoint in tumor immunity that cooperates with CD39 to hydrolyze pro-inflammatory extracellular ATP into immunosuppressive adenosine. While the role of CD73 in immune evasion of solid cancers is well established, its role in leukemia remains unclear. To investigate the role of CD73 in the pathogenesis of chronic lymphocytic leukemia (CLL), Eµ-TCL1 transgenic mice that spontaneously develop CLL were crossed with CD73−/− mice. Disease progression in peripheral blood and spleen, and CLL markers were evaluated by flow cytometry and survival was compared to CD73-proficient Eµ-TCL1 transgenic mice. We observed that CD73 deficiency significantly delayed CLL progression and prolonged survival in Eµ-TCL1 transgenic mice, and was associated with increased accumulation of IFN-γ+ T cells and effector-memory CD8+ T cells. Neutralizing IFN-γ abrogated the survival advantage of CD73-deficient Eµ-TCL1 mice. Intriguingly, the beneficial effects of CD73 deletion were restricted to male mice. In females, CD73 deficiency was uniquely associated with the upregulation of CD39 in normal lymphocytes and sustained high PD-L1 expression on CLL cells. In vitro studies revealed that adenosine signaling via the A2a receptor enhanced PD-L1 expression on Eµ-TCL1-derived CLL cells, and a genomic analysis of human CLL samples found that PD-L1 correlated with adenosine signaling. Our study, thus, identified CD73 as a pro-leukemic immune checkpoint in CLL and uncovered a previously unknown sex bias for the CD73-adenosine pathway. [ABSTRACT FROM AUTHOR]

Simple Summary: Only a limited number of murine mesothelioma cell lines have been developed to date. We sought to expand this number and to characterize the models in detail to enable studying mesothelioma biology in vivo. Two cell lines were identified as showing well-defined mesothelioma biomarkers and being suitable for preclinical use. In the course of our studies, we observed a mixed phenotype of chromosomal instability and microsatellite instability not previously reported in mouse models. Moreover, microsatellite markers were detectable in the plasma of tumor-bearing animals, which potentially can be used as non-invasive biomarkers for early cancer detection and monitoring the effects of interventions. Malignant mesothelioma (MMe) is a rare malignancy originating from the linings of the pleural, peritoneal and pericardial cavities. The best-defined risk factor is exposure to carcinogenic mineral fibers (e.g., asbestos). Genomic studies have revealed that the most frequent genetic lesions in human MMe are mutations in tumor suppressor genes. Several genetically engineered mouse models have been generated by introducing the same genetic lesions found in human MMe. However, most of these models require specialized breeding facilities and long-term exposure of mice to asbestos for MMe development. Thus, an alternative model with high tumor penetrance without asbestos is urgently needed. We characterized an orthotopic model using MMe cells derived from Cdkn2a+/−;Nf2+/− mice chronically injected with asbestos. These MMe cells were tumorigenic upon intraperitoneal injection. Moreover, MMe cells showed mixed chromosome and microsatellite instability, supporting the notion that genomic instability is relevant in MMe pathogenesis. In addition, microsatellite markers were detectable in the plasma of tumor-bearing mice, indicating a potential use for early cancer detection and monitoring the effects of interventions. This orthotopic model with rapid development of MMe without asbestos exposure represents genomic instability and specific molecular targets for therapeutic or preventive interventions to enable preclinical proof of concept for the intervention in an immunocompetent setting. [ABSTRACT FROM AUTHOR]

Simple Summary: Disruption of Bile Acids (BA) regulation with increased BA concentration and modulation or their detergent pro-inflammatory activity has been linked to colorectal cancer (CRC). Farnesoid X Receptor (FXR) is the master regulator of BA homeostasis; FXR is a nuclear receptor that transcriptionally modulates their synthesis, transport and metabolism. In this study, we demonstrated that intestinal FXR activation prevented both inflammation- and genetically-driven colorectal tumorigenesis by modulating BA pool size and composition. This could open new avenues for the therapeutic management of intestinal inflammation and tumorigenesis. The Farnesoid X Receptor (FXR) is the master regulator of Bile Acids (BA) homeostasis orchestrating their synthesis, transport and metabolism. Disruption of BA regulation has been linked to gut-liver axis diseases such as colorectal cancer (CRC). In this study, firstly we examined the role of constitutive activation of intestinal FXR in CRC; then we pre-clinically investigated the therapeutic potential of a diet enriched with a synthetic FXR agonist in two models of CRC (chemically-induced and genetic models). We demonstrated that mice with intestinal constitutive FXR activation are protected from AOM/DSS-induced CRC with a significant reduction of tumor number compared to controls. Furthermore, we evaluated the role of chemical FXR agonism in a DSS model of colitis in wild type (WT) and FXRnull mice. WT mice administered with the FXR activating diet showed less morphological alterations and decreased inflammatory infiltrates compared to controls. The FXR activating diet also protected WT mice from AOM/DSS-induced CRC by reducing tumors' number and size. Finally, we proved that the FXR activating diet prevented spontaneous CRC in APCMin/+ mice via an FXR-dependent modulation of BA homeostasis. Our results demonstrate that intestinal FXR activation prevented both inflammation- and genetically-driven colorectal tumorigenesis by modulating BA pool size and composition. This could open new avenues for the therapeutic management of intestinal inflammation and tumorigenesis. [ABSTRACT FROM AUTHOR]

Silencing the expression of the double homeobox 4 (DUX4) gene offers great potential for the treatment of facioscapulohumeral muscular dystrophy (FSHD). Several research groups have recently reported promising results using systemic antisense therapy in a transgenic small animal model of FSHD, the ACTA1-MCM/FLExDUX4 mouse model. However, the treatment was applied in non-DUX4-induced mice or shortly after DUX4 activation, which resulted in conditions that do not correctly represent the situation in a clinic. Here, we generated progressive FSHD-like pathology in ACTA1-MCM/FLExDUX4 mice and then treated the animals with vivoPMO-PACS4, an antisense compound that efficiently downregulates DUX4. To best mimic the translation of this treatment in clinical settings, the systemic antisense oligonucleotide administration was delayed to 3 weeks after the DUX4 activation so that the pathology was established at the time of the treatment. The chronic administration of vivoPMO-PACS4 for 8 weeks downregulated the DUX4 expression by 60%. Consequently, the treated mice showed an increase by 18% in body-wide muscle mass and 32% in muscle strength, and a reduction in both myofiber central nucleation and muscle fibrosis by up to 29% and 37%, respectively. Our results in a more suitable model of FSHD pathology confirm the efficacy of vivoPMO-PACS4 administration, and highlight the significant benefit provided by the long-term treatment of the disease. [ABSTRACT FROM AUTHOR]

Simple Summary: The main finding reported in this manuscript is that the gain-of-function mutation AKT1E17K is a bona fide oncogene for mammary epithelium, being able to efficiently initiate breast cancer in mice. On the basis of high-molecular-weight cytokeratins expressed by AKT1E17K-derived tumors supported by additional integrative gene expression analysis these tumors resulted similar to human basal-like cancer, phenotypically and molecularly. These results indicate that the AKTE17K strain may represent an appropriate model of human basal-like breast cancer for the identification of novel therapies specific for this type of tumor. The gain-of-function mutation in the pleckstrin homology domain of AKT1 (AKT1E17K) occurs in lung and breast cancer. Through the use of human cellular models and of a AKT1E17K transgenic Cre-inducible murine strain (R26-AKT1E17K mice), we have demonstrated that AKT1E17K is a bona fide oncogene for lung epithelial cells. However, the role of AKT1E17K in breast cancer remains to be determined. Here, we report the generation and the characterization of a MMTV-CRE; R26-AKT1E17K mouse strain that expresses the mutant AKT1E17K allele in the mammary epithelium. We observed that AKT1E17K stimulates the development of mammary tumors classified as ductal adenocarcinoma of medium–high grade and presented a variety of proliferative alterations classified as adenosis with low-to-high grade dysplasia in the mammary epithelium. A subsequent immunohistochemical characterization suggested they were PR−/HER2−/ER+, basal-like and CK8−/CK10−/CK5+/CK14+. We also observed that, in parallel with an increased proliferation rate, tumors expressing mutant AKT1E17K presented an activation of the GSK3/cyclin D1 pathway in the mammary epithelium and cluster significantly with the human basal-like tumors. In conclusion, we demonstrate AKT1E17K is a bona fide oncogene that can initiate tumors at high efficiency in murine mammary epithelium in vivo. [ABSTRACT FROM AUTHOR]

Simple Summary: K14-HPV16 transgenic mice have proved to be a useful model to study the carcinogenic cascade induced by HPV16, the tumor microenvironment and also the epigenetic and genetic factors associated with this type of malignancy. The aim of our study was to evaluate the infiltration of mast cells in two cutaneous regions with different severity of the lesions and to identify potential microRNAs that may regulate mast cell infiltration in this model. We were able to confirm that increased mast cell infiltration is associated with progression of HPV-induced lesions, and that miR-223-3p and miR-125b-5p might be assisting this process via the regulation of mast cell chemotactic proteins. High-risk human papillomavirus (HPV) is the etiologic agent of several types of cancer. Mast cells' role as either a driving or opposing force for cancer progression remains controversial. MicroRNAs are dysregulated in several HPV-induced cancers, and can influence mast cell biology. The aim of this study was to evaluate mast cell infiltration and to identify microRNAs potentially regulating this process. Transgenic male mice (K14-HPV16; HPV+) and matched wild-type mice (HPV−) received 7,12-Dimethylbenz[a]anthracene (DMBA) (or vehicle) over 17 weeks. Following euthanasia, chest skin and ear tissue samples were collected. Mast cell infiltration was evaluated by immunohistochemistry. MicroRNAs associated with mast cell infiltration were identified using bioinformatic tools. MicroRNA and mRNA relative expression was evaluated by RT-qPCR. Immunohistochemistry showed increased mast cell infiltration in HPV+ mice (p < 0.001). DMBA did not have any statistically significant influence on this distribution. Ear tissue of HPV+ mice showed increased mast cell infiltration (p < 0.01) when compared with chest skin samples. Additionally, reduced relative expression of miR-125b-5p (p = 0.008, 2−ΔΔCt = 2.09) and miR-223-3p (p = 0.013, 2−ΔΔCt = 4.42) seems to be associated with mast cell infiltration and increased expression of target gene Cxcl10. These results indicate that HPV16 may increase mast cell infiltration by down-regulating miR-223-3p and miR-125b-5p. [ABSTRACT FROM AUTHOR]

Simple Summary: Ovarian granulosa cell tumors are rare ovarian tumors, with poorly understood etiology. The aim of this study is to define the molecular changes in ovarian granulosa cell tumors. We used a granulosa cell tumor mouse model that contains dysregulated transforming growth factor β signaling. Using RNA-sequencing technology, we identified differentially expressed genes between tumor tissues and normal controls. In addition, comparative analyses have been performed to reveal common genes altered in ovarian granulosa cell tumors between the mouse and the human. This study has revealed the molecular signature of ovarian granulosa cell tumors in a mouse model. The results are potentially important to understand cell signaling events that regulate the development of a class of rare ovarian tumors. Ovarian granulosa cell tumors (GCTs) are rare sex cord-stromal tumors, accounting for ~5% ovarian tumors. The etiology of GCTs remains poorly defined. Genetically engineered mouse models are potentially valuable for understanding the pathogenesis of GCTs. Mice harboring constitutively active TGFβ signaling (TGFBR1-CA) develop ovarian GCTs that phenocopy several hormonal and molecular characteristics of human GCTs. To determine molecular alterations in the ovary upon TGFβ signaling activation, we performed transcriptomic profiling of gene expression associated with GCT development using ovaries from 1-month-old TGFBR1-CA mice and age-matched controls. RNA-sequencing and bioinformatics analysis coupled with the validation of select target genes revealed dysregulations of multiple cellular events and signaling molecules/pathways. The differentially expressed genes are enriched not only for known GCT-related pathways and tumorigenic events but also for signaling events potentially mediated by neuroactive ligand-receptor interaction, relaxin signaling, insulin signaling, and complements in TGFBR1-CA ovaries. Additionally, a comparative analysis of our data in mice with genes dysregulated in human GCTs or granulosa cells overexpressing a mutant FOXL2, the genetic hallmark of adult GCTs, identified some common genes altered in both conditions. In summary, this study has revealed the molecular signature of ovarian GCTs in a mouse model that harbors the constitutive activation of TGFBR1. The findings may be further exploited to understand the pathogenesis of a class of poorly defined ovarian tumors. [ABSTRACT FROM AUTHOR]

Simple Summary: Treatments for castrate (hormone)-resistant prostate cancer (CRPC) remain limited and are not curative. Although the first (and only) FDA-approved vaccine for CRPC (PROVENGE®, Sipuleucel-T) has been shown to improve the overall survival of patients, it is not curative and its cost prevents its widespread use. PROVENGE® induces immunity to prostatic acid phosphatase (PAP), a protein which is highly expressed in prostate cancer (PCa). Herein, we have developed a new PAP-based vaccine for PCa and demonstrated the presence of circulating PAP-specific CD8+ T cells that are responsive to this vaccine in patients with PCa. We have also shown that this new PAP sequence-derived peptide containing a modified amino sequence, in association with a strong adjuvant called CAF®09, induces strong immune responses and cytotoxic potential in a murine HHDII/DR1 transgenic model. Background: Current treatments for castrate (hormone)-resistant prostate cancer (CRPC) remain limited and are not curative, with a median survival from diagnosis of 23 months. The PAP-specific Sipuleucel-T vaccine, which was approved by the FDA in 2010, increases the Overall Survival (OS) by 4 months, but is extremely expensive. We have previously shown that a 15 amino accid (AA) PAP sequence-derived peptide could induce strong immune responses and delay the growth of murine TRAMP-C1 prostate tumors. We have now substituted one amino acid and elongated the sequence to include epitopes predicted to bind to several additional HLA haplotypes. Herein, we present the immunological properties of this 42mer-mutated PAP-derived sequence (MutPAP42mer). Methods: The presence of PAP-135-143 epitope-specific CD8+ T cells in the blood of patients with prostate cancer (PCa) was assessed by flow cytometry using Dextramer™ technology. HHDII/DR1 transgenic mice were immunized with mutated and non-mutated PAP-derived 42mer peptides in the presence of CAF®09 or CpG ODN1826 (TLR-9 agonist) adjuvants. Vaccine-induced immune responses were measured by assessing the proportion and functionality of splenic PAP-specific T cells in vitro. Results: PAP-135-143 epitope-specific CD8+ T cells were detected in the blood of patients with PCa and stimulation of PBMCs from patients with PCa with mutPAP42mer enhanced their capacity to kill human LNCaP PCa target cells expressing PAP. The MutPAP42mer peptide was significantly more immunogenic in HHDII/DR1 mice than the wild type sequence, and immunogenicity was further enhanced when combined with the CAF®09 adjuvant. The vaccine induced secretory (IFNγ and TNFα) and cytotoxic CD8+ T cells and effector memory splenic T cells. Conclusions: The periphery of patients with PCa exhibits immune responsiveness to the MutPAP42mer peptide and immunization of mice induces/expands T cell-driven, wild-type PAP immunity, and therefore, has the potential to drive protective anti-tumor immunity in patients with PCa. [ABSTRACT FROM AUTHOR]

We have recently identified point mutation V336Y in mitoribosomal protein Mrps5 (uS5m) as a mitoribosomal ram (ribosomal ambiguity) mutation conferring error-prone mitochondrial protein synthesis. In vivo in transgenic knock-in animals, homologous mutation V338Y was associated with a discrete phenotype including impaired mitochondrial function, anxiety-related behavioral alterations, enhanced susceptibility to noise-induced hearing damage, and accelerated metabolic aging in muscle. To challenge the postulated link between Mrps5 V338Y-mediated misreading and the in vivo phenotype, we introduced mutation G315R into the mouse Mrps5 gene as Mrps5 G315R is homologous to the established bacterial ram mutation RpsE (uS5) G104R. However, in contrast to bacterial translation, the homologous G → R mutation in mitoribosomal Mrps5 did not affect the accuracy of mitochondrial protein synthesis. Importantly, in the absence of mitochondrial misreading, homozygous mutant MrpS5G315R/G315R mice did not show a phenotype distinct from wild-type animals. [ABSTRACT FROM AUTHOR]

Inverted repeat (IR) DNA sequences compose cruciform structures. Some genetic disorders are the result of genome inversion or translocation by cruciform DNA structures. The present study examined whether exogenous DNA integration into the chromosomes of transgenic animals was related to cruciform DNA structures. Large imperfect cruciform structures were frequently predicted around predestinated transgene integration sites in host genomes of microinjection-based transgenic (Tg) animals (αLA-LPH Tg goat, Akr1A1eGFP/eGFP Tg mouse, and NFκB-Luc Tg mouse) or CRISPR/Cas9 gene-editing (GE) animals (αLA-AP1 GE mouse). Transgene cassettes were imperfectly matched with their predestinated sequences. According to the analyzed data, we proposed a putative model in which the flexible cruciform DNA structures acted as a legible template for DNA integration into linear DNAs or double-strand break (DSB) alleles. To demonstrate this model, artificial inverted repeat knock-in (KI) reporter plasmids were created to analyze the KI rate using the CRISPR/Cas9 system in NIH3T3 cells. Notably, the KI rate of the 5′ homologous arm inverted repeat donor plasmid (5′IR) with the ROSA gRNA group (31.5%) was significantly higher than the knock-in reporter donor plasmid (KIR) with the ROSA gRNA group (21.3%, p < 0.05). However, the KI rate of the 3′ inverted terminal repeat/inverted repeat donor plasmid (3′ITRIR) group was not different from the KIR group (23.0% vs. 22.0%). These results demonstrated that the legibility of the sequence with the cruciform DNA existing in the transgene promoted homologous recombination (HR) with a higher KI rate. Our findings suggest that flexible cruciform DNAs folded by IR sequences improve the legibility and accelerate DNA 3′-overhang integration into the host genome via homologous recombination machinery. [ABSTRACT FROM AUTHOR]

The accumulation of iron may contribute to Alzheimer's disease (AD) and other tauopathies. The iron chelator desferrioxamine slows disease progression in AD patients. However, desferrioxamine requires injection, which is inconvenient and may hinder compliance. We therefore tested an oral iron chelator, desferasirox (Exjade), in transgenic animal models. Tg2576 mice overexpress the mutant human APP protein and produce the Aβ peptide. JNPL3 mice (Tau/Tau) overexpress the mutant human tau protein. Crossing these produced APP/Tau mice, overexpressing both APP and tau. Treating the three models with 1.6 mg deferasirox thrice weekly from age 8 to 14 months did not affect memory as measured by contextual fear conditioning or motor function as measured by rotarod, but tended to decrease hyperphosphorylated tau as measured by AT8 immunohistochemistry and immunoblotting. Deferasirox might act by decreasing iron, which aggregates tau, or directly binding tau to inhibit aggregation. [ABSTRACT FROM AUTHOR]

There is a need for improved in vitro models of inherited cardiac diseases to better understand basic cellular and molecular mechanisms and advance drug development. Most of these diseases are associated with arrhythmias, as a result of mutations in ion channel or ion channel-modulatory proteins. Thus far, the electrophysiological phenotype of these mutations has been typically studied using transgenic animal models and heterologous expression systems. Although they have played a major role in advancing the understanding of the pathophysiology of arrhythmogenesis, more physiological and predictive preclinical models are necessary to optimize the treatment strategy for individual patients. Human induced pluripotent stem cell-derived cardiomyocytes (hiPSC-CMs) have generated much interest as an alternative tool to model arrhythmogenic diseases. They provide a unique opportunity to recapitulate the native-like environment required for mutated proteins to reproduce the human cellular disease phenotype. However, it is also important to recognize the limitations of this technology, specifically their fetal electrophysiological phenotype, which differentiates them from adult human myocytes. In this review, we provide an overview of the major inherited arrhythmogenic cardiac diseases modeled using hiPSC-CMs and for which the cellular disease phenotype has been somewhat characterized. [ABSTRACT FROM AUTHOR]

Pancreatic Ductal Adenocarcinoma (PDA) is an almost incurable radio- and chemo-resistant tumor, and its microenvironment is characterized by a strong desmoplastic reaction associated with a significant infiltration of T regulatory lymphocytes and myeloid-derived suppressor cells (Tregs, MDSC). Investigating immunological targets has identified a number of metabolic and cytoskeletal related molecules, which are typically recognized by circulating antibodies. Among these molecules we have investigated alpha-enolase (ENO1), a glycolytic enzyme that also acts a plasminogen receptor. ENO1 is also recognized by T cells in PDA patients, so we developed a DNA vaccine that targets ENO1. This efficiently induces many immunological processes (antibody formation and complement-dependent cytotoxicity (CDC)-mediated tumor killing, infiltration of effector T cells, reduction of infiltration of myeloid and Treg suppressor cells), which significantly increase the survival of genetically engineered mice that spontaneously develop pancreatic cancer. Although promising, the ENO1 DNA vaccine does not completely eradicate the tumor, which, after an initial growth inhibition, returns to proliferate again, especially when Tregs and MDSC ensue in the tumor mass. This led us to develop possible strategies for combinatorial treatments aimed to broaden and sustain the antitumor immune response elicited by DNA vaccination. Based on the data we have obtained in recent years, this review will discuss the biological bases of possible combinatorial treatments (chemotherapy, PI3K inhibitors, tumor-associated macrophages, ENO1 inhibitors) that could be effective in amplifying the response induced by the immune vaccination in PDA. [ABSTRACT FROM AUTHOR]

The rapid development of genome modification technology has provided many great benefits in diverse areas of research and industry. Genome modification technologies have also been actively used in a variety of research areas and fields of industry in avian species. Transgenic technologies such as lentiviral systems and piggyBac transposition have been used to produce transgenic birds for diverse purposes. In recent years, newly developed programmable genome editing tools such as transcription activator-like effector nuclease (TALEN) and clustered regularly interspaced short palindromic repeats (CRISPR)/CRISPR-associated protein 9 (CRISPR/Cas9) have also been successfully adopted in avian systems with primordial germ cell (PGC)-mediated genome modification. These genome modification technologies are expected to be applied to practical uses beyond system development itself. The technologies could be used to enhance economic traits in poultry such as acquiring a disease resistance or producing functional proteins in eggs. Furthermore, novel avian models of human diseases or embryonic development could also be established for research purposes. In this review, we discuss diverse genome modification technologies used in avian species, and future applications of avian biotechnology. [ABSTRACT FROM AUTHOR]