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Abstract Background Alzheimer's disease (AD) is a devastating neurodegenerative disorder that affects a large and growing number of elderly individuals. In addition to idiopathic disease, AD is also associated with autosomal dominant inheritance, which causes a familial form of AD (FAD). Some instances of FAD have been linked to mutations in the β-amyloid protein precursor (APP). Although there are numerous mouse AD models available, few rat AD models, which have several advantages over mice, have been generated. Results Fischer 344 rats expressing human APP driven by the ubiquitin-C promoter were generated via lentiviral vector infection of Fischer 344 zygotes. We generated two separate APP-transgenic rat lines, APP21 and APP31. Serum levels of human amyloid-beta (Aβ)40 were 298 pg/ml for hemizygous and 486 pg/ml for homozygous APP21 animals. Serum Aβ42 levels in APP21 homozygous rats were 135 pg/ml. Immunohistochemistry in brain showed that the human APP transgene was expressed in neurons, but not in glial cells. These findings were consistent with independent examination of enhanced green fluorescent protein (eGFP) in the brains of eGFP-transgenic rats. APP21 and APP31 rats expressed 7.5- and 3-times more APP mRNA, respectively, than did wild-type rats. Northern blots showed that the human APP transgene, driven by the ubiquitin-C promoter, is expressed significantly more in brain, kidney and lung compared to heart and liver. A similar expression pattern was also seen for the endogenous rat APP. The unexpected similarity in the tissue-specific expression patterns of endogenous rat APP and transgenic human APP mRNAs suggests regulatory elements within the cDNA sequence of APP. Conclusion This manuscript describes the generation of APP-transgenic inbred Fischer 344 rats. These are the first human AD model rat lines generated by lentiviral infection. The APP21 rat line expresses high levels of human APP and could be a useful model for AD. Tissue-specific expression in the two transgenic rat lines and in wild-type rats contradicts our current understanding of APP gene regulation. Determination of the elements that are responsible for tissue-specific expression of APP may enable new treatment options for AD.

Background: Alzheimer's disease (AD) is a devastating neurodegenerative disorder that affects a large and growing number of elderly individuals. In addition to idiopathic disease, AD is also associated with autosomal dominant inheritance, which causes a familial form of AD (FAD). Some instances of FAD have been linked to mutations in the beta-amyloid protein precursor (APP). Although there are numerous mouse AD models available, few rat AD models, which have several advantages over mice, have been generated.Results: Fischer 344 rats expressing human APP driven by the ubiquitin-C promoter were generated via lentiviral vector infection of Fischer 344 zygotes. We generated two separate APP-transgenic rat lines, APP21 and APP31. Serum levels of human amyloid-beta (Abeta)40 were 298 pg/ml for hemizygous and 486 pg/ml for homozygous APP21 animals. Serum Abeta42 levels in APP21 homozygous rats were 135 pg/ml. Immunohistochemistry in brain showed that the human APP transgene was expressed in neurons, but not in glial cells. These findings were consistent with independent examination of enhanced green fluorescent protein (eGFP) in the brains of eGFP-transgenic rats. APP21 and APP31 rats expressed 7.5- and 3-times more APP mRNA, respectively, than did wild-type rats. Northern blots showed that the human APP transgene, driven by the ubiquitin-C promoter, is expressed significantly more in brain, kidney and lung compared to heart and liver. A similar expression pattern was also seen for the endogenous rat APP. The unexpected similarity in the tissue-specific expression patterns of endogenous rat APP and transgenic human APP mRNAs suggests regulatory elements within the cDNA sequence of APP.Conclusion: This manuscript describes the generation of APP-transgenic inbred Fischer 344 rats. These are the first human AD model rat lines generated by lentiviral infection. The APP21 rat line expresses high levels of human APP and could be a useful model for AD. Tissue-specific expression in the two transgenic rat lines and in wild-type rats contradicts our current understanding of APP gene regulation. Determination of the elements that are responsible for tissue-specific expression of APP may enable new treatment options for AD. [ABSTRACT FROM AUTHOR]

Unlike mild DNA damage exposure, DNA damage repair (DDR) is reported to be ineffective in full-grown mammalian oocytes exposed to moderate or severe DNA damage. The underlying mechanisms of this weakened DDR are unknown. Here, we show that moderate DNA damage in full-grown oocytes leads to aneuploidy. Our data reveal that DNA-damaged oocytes have an altered, closed, chromatin state, and suggest that the failure to repair damaged DNA could be due to the inability of DDR proteins to access damaged loci. Our data also demonstrate that, unlike somatic cells, mouse and porcine oocytes fail to activate autophagy in response to DNA double-strand break-inducing treatment, which we suggest may be the cause of the altered chromatin conformation and inefficient DDR. Importantly, autophagy activity is further reduced in maternally aged oocytes (which harbor severe DNA damage), and its induction is correlated with reduced DNA damage in maternally aged oocytes. Our findings provide evidence that reduced autophagy activation contributes to weakened DDR in oocytes, especially in those from aged females, offering new possibilities to improve assisted reproductive therapy in women with compromised oocyte quality., (© 2024. The Author(s).)

Now in its 25th year, the Mutant Mouse Resource and Research Center (MMRRC) consortium continues to serve the United States and international biomedical scientific community as a public repository and distribution archive of laboratory mouse models of human disease for research. Supported by the National Institutes of Health (NIH), the MMRRC consists of 4 regionally distributed and dedicated vivaria, offices, and specialized laboratory facilities and an Informatics Coordination and Service Center (ICSC). The overarching purpose of the MMRRC is to facilitate groundbreaking biomedical research by offering an extensive repertoire of mutant mice that are essential for advancing the understanding of human physiology and disease. The function of the MMRRC is to identify, acquire, evaluate, characterize, cryopreserve, and distribute mutant mouse strains to qualified biomedical investigators around the nation and the globe. Mouse strains accepted from the research community are held to the highest scientific standards to optimize reproducibility and enhance scientific rigor and transparency. All submitted strains are thoroughly reviewed, documented, and validated using extensive scientific quality control measures. In addition, the MMRRC conducts resource-related research on cryopreservation, mouse genetics, environmental conditions, and other topics that enhance operations of the MMRRC. Today, the MMRRC maintains an archive of mice, cryopreserved embryos and sperm, embryonic stem (ES) cell lines, and murine hybridomas for nearly 65,000 alleles. Since its inception, the MMRRC has fulfilled more than 20,000 orders from 13,651 scientists at 8441 institutions worldwide. The MMRRC also provides numerous services to assist researchers, including scientific consultation, technical assistance, genetic assays, microbiome analysis, analytical phenotyping, pathology, cryorecovery, husbandry, breeding and colony management, infectious disease surveillance, and disease modeling. The ICSC coordinates MMRRC operations, interacts with researchers, and manages the website (mmrrc.org) and online catalogue. Researchers benefit from an expansive list of well-defined mouse models of disease that meet the highest scientific standards while submitting investigators benefit by having their mouse strains cryopreserved, protected, and distributed in compliance with NIH policies., (© 2024. The Author(s).)

Conventional, brightfield-microscopic semen analysis provides important baseline information about sperm quality of an individual; however, it falls short of identifying subtle subcellular and molecular defects in cohorts of "bad," defective human and animal spermatozoa with seemingly normal phenotypes. To bridge this gap, it is desirable to increase the precision of andrological evaluation in humans and livestock animals by pursuing advanced biomarker-based imaging methods. This review, spiced up with occasional classic movie references but seriously scholastic at the same time, focuses mainly on the biomarkers of altered male germ cell proteostasis resulting in post-testicular carryovers of proteins associated with ubiquitin-proteasome system. Also addressed are sperm redox homeostasis, epididymal sperm maturation, sperm-seminal plasma interactions, and sperm surface glycosylation. Zinc ion homeostasis-associated biomarkers and sperm-borne components, including the elements of neurodegenerative pathways such as Huntington and Alzheimer disease, are discussed. Such spectrum of biomarkers, imaged by highly specific vital fluorescent molecular probes, lectins, and antibodies, reveals both obvious and subtle defects of sperm chromatin, deoxyribonucleic acid, and accessory structures of the sperm head and tail. Introduction of next-generation image-based flow cytometry into research and clinical andrology will soon enable the incorporation of machine and deep learning algorithms with the end point of developing simple, label-free methods for clinical diagnostics and high-throughput phenotyping of spermatozoa in humans and economically important livestock animals., (© The Author(s) 2024. Published by Oxford University Press on behalf of Society for the Study of Reproduction.)

-In this paper, we investigate a microfluidic based sensing device for cell membrane permeability measurements in real time with applications in rapid assessment of red blood cell (RBC) quality at the individual cell level. The microfluidic chip was designed with unique abilities to line up the RBCs in the centerline of the microchannel using positive dielectrophoresis (p-DEP) forces, rapid mixing of RBCs with various media (e.g. containing permeating or nonpermeating solutes) injected from different inlets to achieve high mixing efficiency. The chip detects the impedance values of the RBCs within 0.19 s from the start of mixing with other media, at ten electrodes along the length of the channel and enables time series measurements of volume change of individual cell caused by cell osmosis in anisosmotic fluids over a 0.8 s postmixing timespan. This technique enables estimating water permeability of individual cell accurately. Here we first present confirmation of a linear voltage-diameter relationship in polystyrene bead standards. Next, we show that under equilibrium conditions, the voltage-volume relationship in rat red blood cells (RBCs) is linear, corresponding to previously published Boyle van 't Hoff plots. Using rat cells as a model for human, we present the first measurement of water permeability in individual red blood cells and confirm that these data align with previously published population level values for human RBC. Finally, we present preliminary evidence for possible application of our device to identify individual RBCs infected with Plasmodium falciparum malaria parasites. Future developments using this device will address the use of whole blood with non-homogenous cell populations, a task currently performed by clinical Coulter counters., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023. Published by Elsevier B.V.)

Cryopreservation of epididymal sperm collected after euthanasia is a common method to preserve and distribute valuable mouse models worldwide. However, the euthanasia method used prior to sperm collection must not adversely affect sperm quality. The most common method of euthanasia in mice is CO₂ asphyxiation, but its effect on the quality of sperm collected postmortem is largely unknown. The objective of this study was to determine the effects of CO₂ euthanasia of C57BL/6 mice on both freshly recovered sperm and sperm subjected to freezing and thawing. First, sperm concentration, progressive motility, curvilineal velocity (VCL), average path velocity (VAP), and progressive velocity (VSL) were analyzed for mice euthanized by cervical dislocation (CD), high flow CO₂ (100%), or low flow CO₂ (30%) displacement/minute, respectively. Then, in-vitro fertilization and embryonic development rates were determined using frozen-thawed sperm from each euthanasia method. Neither fresh nor frozen-thawed sperm showed significant differences in sperm concentration, progressive motility, VAP, or VCL when compared to CD and CO₂ groups. However, frozen-thawed sperm collected from CD mice had higher VCL values than did those collected from the low flow mice ( P = 0.039). VCL was not different in fresh or frozen-thawed sperm collected after mouse euthanasia by CD as compared with high flow CO₂ or by high flow as compared with low flow CO₂. Frozen-thawed sperm showed no differences among the 3 euthanasia groups for fertilization ( P = 0.452) or blastocyst development rates ( P = 0.298). The results indicate that CO₂ euthanasia can be used as an alternative to CD to obtain optimal quality mouse sperm for cryopreservation while remaining compliant with welfare requirements.

Ischemic stroke often co-occurs with Alzheimer's disease (AD) leading to a worsened clinical outcome. Neuroinflammation is a critical process implicated in AD and ischemic pathology, associated with cognitive decline. We sought to investigate the combined effects of ischemic stroke induced by endothelin-1 injection in two AD rat models, using motor function, memory and microglial inflammation in the basal forebrain and striatum as readouts. In addition, we sought to determine the effectiveness of the antioxidant biologic CAT-SKL in one of the models. The early AD model employed the bilateral intracerebroventricular injections of the toxic β-amyloid peptide Aβ 25-35 , the prodromal AD model used the transgenic Fischer 344 rat overexpressing a pathological mutant human amyloid precursor protein. Motor function was assessed using a cylinder, modified sticky tape and beam-walk tasks; learning and memory were tested in the Morris water maze. Microglial activation was examined using immunohistochemistry. Aβ 25-35 toxicity and stroke combination greatly increased microglial inflammation in the basal forebrain. Prodromal AD-pathology coupled with ischemia in the transgenic rat resulted in a greater microgliosis in the striatum. Combined transgenic rats showed balance alterations, comorbid Aβ 25-35 rats showed a transient sensorimotor deficit, and both demonstrated spatial reference memory deficit. CAT-SKL treatment ameliorated memory impairment and basal forebrain microgliosis in Aβ 25-35 rats with stroke. Our results suggest that neuroinflammation could be one of the early processes underlying the interaction of AD with stroke and contributing to the cognitive impairment, and that therapies such as antioxidant CAT-SKL could be a potential therapeutic strategy., (Copyright © 2022 The Author(s). Published by Elsevier Ltd.. All rights reserved.)

Neuroinflammation and behavioural inflexibility are both common in late adulthood but far more profound in Alzheimer disease (AD). To investigate the relationship between ageing, AD, neuroinflammation, and behavioural flexibility, male wild-type Fischer 344 (Wt) and the transgenic APP21 (TgAPP21) rats were aged to 4, 8, 13, and 22 months and evaluated for neuroinflammation and cognitive impairment. TgAPP21 rats overexpress a pathogenic variant of the human amyloid precursor protein (hAPP; Swedish and Indiana mutations) but do not spontaneously develop overt pathology related to AD. In both genotypes, learning and memory were similarly impaired in older rats. However, at 8 months of age, TgAPP21 rats demonstrated behavioural inflexibility in set shifting, reversal, and the Morris water maze, while Wt rats showed inflexibility at 13 and 22 months of age. This early inflexibility in TgAPP21 rats was accompanied by a precocious increase in microglia activation within the corpus callosum; 8- and 13-month-old TgAPP21 rats had similar levels of microglia activation as 13- and 22-month-old Wt rats, respectively. However, while neuroinflammation within the white matter continued to progress with age, behavioural inflexibility peaked in 8-month-old TgAPP21 rats; in older TgAPP21 rats, memory and learning impairments masked inflexibility. These findings suggest that the behavioural inflexibility and white matter inflammation seen in normal ageing are accelerated in AD and may precede impairments of learning and memory., (© 2021. The Author(s), under exclusive licence to Springer Science+Business Media, LLC, part of Springer Nature.)

Genetic modifications in combination with highly sophisticated assisted reproductive technologies such as in vitro oocyte maturation and development, in vitro fertilization, intracytoplasmic sperm injection, and in vitro embryo culture have opened many research avenues and treatment options for both animals and humans. The number of genetically modified (GM) rodent strains increased considerably during the last several decades, and their numbers are expected to increase due to efficient gene editing technologies including the CRISPR/Cas9. Rodent ovarian tissues (OT) cryopreservation and transplantation procedures have several applications in biomedical field: they provide a fertility restoration option for GM rodent strains in some circumstances. They also serve as models to investigate OT cryopreservation as potential alternatives for human infertility patients as well as other domestic and wildlife species for the development of improved cryopreservation and subsequent transplantation strategies. The modeling studies enable determining effective cryoprotective agents (CPA), CPA and water permeability kinetics, and cooling and warming rates during the development of OT cryopreservation procedures. Furthermore, rodent models are extremely useful for determining post-thaw OT graft sites as well as potential medical interventions in an effort to expedite angiogenesis and inhibit inflammatory/immune response, OT longevity, and follicular integrity. Here we describe methodologies for rodent OT cryopreservation and potential transplantation sites for frozen-thawed rat and mouse OT.

Germplasm cryobanking of transgenic rodent models is a valuable tool for protecting important genotypes from genetic drift, genetic contamination, and loss of breeding colonies due to disease or catastrophic disasters to the housing facilities as well as avoiding stress associated with domestic and international live animal shipment. Furthermore, cryopreservation of germplasm enhances management efficiencies by saving animal room space, reducing workload for staff, reducing cost of maintaining live animals, reducing the number of animals used to maintain a breeding colony, and facilitating transportation of genetics by allowing distribution of frozen germplasm rather than live animals which also reduces the risk of transfer of pathogens between facilities. Thus, effective long-term preservation methods of mouse spermatozoa are critical for future reconstitution of scientifically important mouse strains used for biomedical research.

Hypertension is recognized as a risk factor for Alzheimer disease, but the causal link remains undetermined. Although astrocytes and microglia play an important role in maintaining the neurovascular unit, astrocytes and microglia have been understudied in comorbid models of hypertension and Alzheimer disease. In this study, male transgenic Fischer 344 rats (TgAPP21) overexpressing a pathogenic human amyloid precursor protein received 8 weeks of Angiotensin II infusion to increase blood pressure, and the rats were evaluated for astrocytosis, microgliosis, and cognitive function. A linear relationship between astrocytosis and blood pressure was observed in the corpus callosum and cingulum of wildtype rats, with hypertensive wildtype rats matching the elevated baseline astrocytosis seen in normotensive transgenic rats. In contrast, hypertensive transgenic rats did not demonstrate a further increase of astrocytosis, suggesting a deficient response. Angiotensin II infusion did not affect activation of microglia, which were elevated in the white matter and hippocampus of transgenic rats. Angiotensin II infusion did impair both wildtype and transgenic rats' executive functions in the Morris Water Maze. These results present important implications for the interaction between hypertension and pathogenic human amyloid precursor protein expression, as Angiotensin II infusion produced cognitive impairments in both genotypes, but transgenic rats were additionally impaired in developing a normal astrocytic response to elevated blood pressure., (Copyright © 2020 Levit, Cheng, Hough, Liu, Agca, Agca, Hachinski and Whitehead.)