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Introduction Traumatic brain injury is a major public health concern. Among patients with severe traumatic brain injury, epidural hemorrhage is known to swiftly lead to brain herniation and death unless there is emergent neurosurgical intervention. However, immediate neurosurgeon availability is frequently a problem outside of level I trauma centers. In this context, the authors desired to test a novel device for the emergent management of life-threatening epidural hemorrhage. A review of existing animal models determined that all were inadequate for this purpose, as they were found to be either inappropriate or obsolete. Here, we describe the development of a new epidural hemorrhage model in swine (Sus scrofa, 18-26 kg) ideal for translational device testing. Materials and Methods Vascular access was achieved using an ultrasound-guided percutaneous Seldinger catheter-over-wire technique with 5 Fr catheters placed in the bilateral carotid arteries, for continuous blood pressure and to allow for withdrawal of blood for creation of epidural hemorrhage. To simulate an actively bleeding and life-threatening epidural hemorrhage, unadulterated autologous blood was infused from the vascular access point into the epidural space. To be useful for this application and clinical scenario, brain death needed to occur after the planned intervention time but before the end of the protocol period (if no intervention took place). An iterative approach to model development determined that this could be achieved with an initial infusion rate of 1.0 mL/min, slowed to 0.5 mL/min after the first 10 minutes, paired with an intervention time at 15 minutes. All experiments were performed at Oregon Health & Science University, an Association for Assessment and Accreditation of Laboratory Animal Care accredited facility. Oregon Health & Science University’s Institutional Animal Care and Use Committee, as well as the United States Army Animal Care and Use Review Office, reviewed and approved this protocol before the initiation of experiments (respectively, protocol numbers IP00002901 and 18116010.e001). Results The final developed model allows for the infusion of a known volume of autologous, unadulterated blood directly into the epidural space, without the use of a balloon or other restricting membranes, and is rapidly fatal in the absence of intervention. Conclusions This animal model is the first to mirror the expected clinical course of epidural hemorrhage in a physiologically relevant manner, while allowing translational testing of emergency devices. This model successfully allowed the initial testing of a novel interventional device for the emergent management of epidural hemorrhage that was designed for use in the absence of traditional neurosurgical capabilities.

Magnetic resonance imaging (MRI) is a clinically relevant, real-time imaging modality that is frequently utilized to assess stroke type and severity. However, specific MRI biomarkers that can be used to predict long-term functional recovery are still a critical need. Consequently, the present study sought to examine the prognostic value of commonly utilized MRI parameters to predict functional outcomes in a porcine model of ischemic stroke. Stroke was induced via permanent middle cerebral artery occlusion. At 24 hours post-stroke, MRI analysis revealed focal ischemic lesions, decreased diffusivity, hemispheric swelling, and white matter degradation. Functional deficits including behavioral abnormalities in open field and novel object exploration as well as spatiotemporal gait impairments were observed at 4 weeks post-stroke. Gaussian graphical models identified specific MRI outputs and functional recovery variables, including white matter integrity and gait performance, that exhibited strong conditional dependencies. Canonical correlation analysis revealed a prognostic relationship between lesion volume and white matter integrity and novel object exploration and gait performance. Consequently, these analyses may also have the potential of predicting patient recovery at chronic time points as pigs and humans share many anatomical similarities (e.g., white matter composition) that have proven to be critical in ischemic stroke pathophysiology. The study was approved by the University of Georgia (UGA) Institutional Animal Care and Use Committee (IACUC; Protocol Number: A2014-07-021-Y3-A11 and 2018-01-029-Y1-A5) on November 22, 2017.

Intraspinal inflammatory and immune responses are considered to play central roles in the pathological development of spinal cord injury. This study aimed to decipher the dynamics of systemic immune responses, initiated by spinal cord injury. The spinal cord in mice was completely transected at T8. Changes in the in vivo inflammatory response, between the acute and subacute stages, were observed. A rapid decrease in C-reactive protein levels, circulating leukocytes and lymphocytes, spleen-derived CD4+ interferon-γ+ T-helper cells, and inflammatory cytokines, and a marked increase in neutrophils, monocytes, and CD4+ CD25+ FOXP3+ regulatory T-cells were observed during the acute phase. These systemic immune alterations were gradually restored to basal levels during the sub-acute phase. During the acute phase of spinal cord injury, systemic immune cells and factors showed significant inhibition; however, this inhibition was transient, and the indicators of these serious disorders gradually returned to baseline levels during the subacute phase. All experiments were performed in accordance with the institutional animal care guidelines, approved by the Institutional Animal Care and Use Committee of Experimental Animal Center of Drum Tower Hospital, China (approval No. 2019AE01040) on June 25, 2019.

Is it ethically permissible to acquire biological materials from a slaughterhouse for biomedical research? This essay examines this question, using a recent, high-profile research program as a case study. Using roughly 300 decapitated pig heads acquired from a slaughterhouse, researchers reperfused the animals' brains and observed a variety of cellular and molecular activities. The study was exempted from review and oversight by the Institutional Animal Care and Use Committee (IACUC), on the grounds that the animals were already dead. This essay argues that the IACUC was mistaken in exempting the study from their oversight, and furthermore, that acquiring animals or their parts from a slaughterhouse for research purposes is unethical and should not occur again. Examination of the study within a broader societal context helps to illuminate why each of us has an ethical obligation to do what we can to abolish the vicious and cruel treatment endured by billions of animals every year on factory farms and in slaughterhouses.

Apoptosis is an important programmed cell death process involved in ischemia/reperfusion injury. MicroRNAs are considered to play an important role in the molecular mechanism underlying the regulation of cerebral ischemia and reperfusion injury. However, whether miR-670 can regulate cell growth and death in cerebral ischemia/reperfusion and the underlying mechanism are poorly understood. In this study, we established mouse models of transient middle artery occlusion and Neuro 2a cell models of oxygen-glucose deprivation and reoxygenation to investigate the potential molecular mechanism by which miR-670 exhibits its effects during cerebral ischemia/reperfusion injury both in vitro and in vivo. Our results showed that after ischemia/reperfusion injury, miR-670 expression was obviously increased. After miR-670 expression was inhibited with an miR-670 antagomir, cerebral ischemia/reperfusion injury-induced neuronal death was obviously reduced. When miR-670 overexpression was induced by an miR-670 agomir, neuronal apoptosis was increased. In addition, we also found that miR-670 could promote Yap degradation via phosphorylation and worsen neuronal apoptosis and neurological deficits. Inhibition of miR-670 reduced neurological impairments after cerebral ischemia/reperfusion injury. These results suggest that microRNA-670 aggravates cerebral ischemia/reperfusion injury through the Yap pathway, which may be a potential target for treatment of cerebral ischemia/reperfusion injury. The present study was approved by the Institutional Animal Care and Use Committee of China Medical University on February 27, 2017 (IRB No. 2017PS035K).

Sunitinib has been used as the first-line therapy to suppress the metastatic clear cell Renal Cell Carcinoma (ccRCC) as it could function via suppressing the tumor growth and angiogenesis. Yet most ccRCC tumors may still regrow due to the development of sunitinibresistance, and detailed mechanisms remain unclear. Here, we found the estrogen receptor β (ERβ) in RCC cells might function via increasing the endothelial cell tube formation to alter the sensitivity to sunitinib treatment. Mechanism dissection revealed that ERβ could function via transcriptional regulation with direct binding to the promoter to increase the cytokine Angiopoietin-2 expression in the ccRCC cells. The up-regulated Angiopoietin-2 could then increase the Tie-2 phosphorylation in the endothelial Huvec cells to promote the angiogenesis tube formation that results in increasing the endothelial cell resistance to sunitinib treatment. Preclinical study using an in vivo mouse RCC model also confirm in vitro data. Targeting this newly identified ERβ/Angiopoietin2/Tie-2 signaling-increased angiogenesis tube formation with the FDAapproved anti-estrogen, ICI 182,780 (Faslodex) may help in the development of a novel combined therapy with sunitinib to better suppress the ccRCC progression. Funding Statement: This work was partially supported by the URMC Urology research fund and the George Whipple Professorship Endowment. Declaration of Interests: The authors declare no potential conflicts of interest. Ethics Approval Statement: The Institutional Animal Care and Use Committee (IACUC) review board at University of Rochester Medical Center approved the studies.

Background: Hypothalamic astrocytes are the important contributors that activate gonadotropin-releasing hormone (GnRH) neurons and promote GnRH/LH (luteinizing hormone) surge. However, the underlying roles and mechanisms of astrocytes during the early age reproductive decline remain unclear. Methods: Immunofluorescence and immunohistochemical staining were applied to analyze the expression of p16, γ-H2AX, SA-β-Gal and peroxidase. Animal ovariectomized models and cell models receiving repeated estradiol intervention were adopted to investigate estradiol-induced astrocyte senescence. High-performance liquid chromatography (HPLC) was employed to detect the estradiol metabolites and progesterone. qPCR and western blot were performed to analyze the PKA-CYPs pathway, growth factors and proinflammatory cytokines. Findings: Intact middle-aged female mice, astrocytes within the hypothalamic RP3V accumulate senescence-related markers with increasing age. Molecular analysis revealed that ovarian estradiol activated PKA and up-regulated CYPs (cytochrome p450) expression, metabolizing estradiol into 2-OHE2 and 4-OHE2 and mediating the senescence of astrocytes in RP3V. Of note, the progesterone synthesis and the ability to promote GnRH release were significantly reduced in middle-aged mice. Besides, the expression of growth factors decreased and the mRNA levels of proinflammatory cytokines significantly increased in the aging astrocytes. Interpretation: The findings confirm that ovarian estradiol induces the senescence of hypothalamic astrocytes. The senescent astrocytes compromises the function for regulating progesterone synthesis and GnRH secretion, may be contributes to aging-related declines in female reproductive function. Funding Statement: The study was supported by National Natural Science Foundation of China to Jing Zhang (No.81671400) and Fujian Province Health and Family Planning Youth Research Project to Xiaoman Dai(No.2018-1-45). Declaration of Interests: All authors have no potential conflicts of interests to declare, financial or otherwise Ethics Approval Statement: All protocols and procedures used in these studies involving animals were approved by the Institutional Animal Care and Use Committee of Fujian Medical University and in compliance with NIH’s Guidelines for the Care and Use of Laboratory Animals (NIH Publication No. 85-23 Rev. 1985). This article does not contain any studies with human participants performed by any of the authors.

Ensuring appropriate review, approval, and oversight of research involving animals becomes increasingly complex when researchers collaborate across multiple sites. In these situations, it is important that the division of responsibilities is clear and that all involved parties share a common understanding. The National Institutes of Health Office of Laboratory Animal Welfare and the United States Department of Agriculture Animal Plant Health Inspection Service require an Institutional Animal Care and Use Committee (IACUC) to review the care and use of animals in research, and both agree that it is acceptable for one IACUC to review the work taking place at multiple institutions. With this in mind, several Harvard-affiliated hospitals and academic centers developed the Master Reciprocal Institutional Agreement for Animal Care and Use (Master IACUC Agreement) to support collaboration, decrease administrative burden, increase efficiencies, reduce duplicative efforts, and ensure appropriate protections for animals used in research. Locally, the Master IACUC Agreement has fostered greater collaboration and exchange while ensuring appropriate review and oversight of research involving animals. As multisite animal protocols become more prevalent, this Agreement could provide a model for a distributed, national network of IACUC reliance.

Background: Hyperbilirubinemia may increase the risk of Alzheimer's disease (AD) but its mechanistic role in AD pathogenesis remains obscure. Methods: Three animal models and primary hippocampal cells were used to explore in this study: (1) acute bilirubin treatment; (2) lateral ventricle injection; (3) long term observation. Results: Three days exposure to bilirubin in newborn rats could induce AD-like pathological changes in late life, including tau protein hyperphosphorylation at multiple sites, increased Aβ production in brain tissues, and spatial learning and memory injury. Bilirubin activated the activities of several protein kinases (GSK-3β, CDK5 and JNK), which were positively correlated with hyperphosphorylated tau; simultaneously increased the expression of APP γ-secretase PS2 and decreased the expression of α-secretase ADAM17, which were positively correlated with Aβ production. The above results were well replicated in primary hippocampal cells. Conclusions: Bilirubin may be an upstream activator to induce AD, and bilirubin encephalopathy is an AD-like disease. Funding Statement: The study was supported in part by the National High Technology Research and Development Program of China (2006AA02Z4B5); the National Natural Science Foundation of China(81170608, 81570278, 81602427, 81600089, 81670156); the Science and Technology Program of Guangzhou (201707010046, 201604020001); The National Funds for Developing Local Colleges and Universities (B16056001); Natural Science Foundation research team of Guangdong Province (2018B030312001); Guangzhou Education System (1201610014, 1201620494); Natural Science Foundation of Guangdong Province (2017A030313662). Declaration of Interests: All authors declare that there are no conflicts of interest. Ethics Approval Statement: The protocol for the care and use of all animals in this study was in accordance with the Guangdong Animal Center for the ethical treatment of animals and approved by the Institutional Animal Care and Use Committee of Guangzhou Medical University (SYXK2016-0168, Guangzhou, China).

Spinal cord injury (SCI) leads to disruption of the blood-spinal cord barrier, hemorrhage and tissue edema, which impair blood circulation and induce ischemia. Angiogenesis after SCI is an important step in the repair of damaged tissues, and its extent strongly correlates with neural regeneration. Various biomaterials have been developed to promote angiogenesis signaling pathways, and angiogenic self-assembling peptides are useful for producing diverse supramolecular structures with tunable functionality. RADA16 (Ac-RARADADARARADADA-NH2), which forms nanofiber networks under physiological conditions, is a self-assembling peptide that can provide mechanical support for tissue regeneration and reportedly has diverse roles in wound healing. In this study, we applied an injectable form of RADA16 with or without a neuropeptide, substance P, to the contused spinal cord in rats and examined angiogenesis within the damaged spinal cord and subsequent functional improvement. Histological and immunohistochemical analyses revealed that the inflammatory cell population in the lesion cavity was decreased, whereas the vessel number and density around the damaged spinal cord were increased, and neurofilaments within the lesion cavity were increased in SCI rats that received RADA16 and RADA16 with substance P (RADA16/SP group). Moreover, real-time PCR of the damaged spinal cord showed that IL-10 expression was increased, and locomotor function (assessed by the Basso, Beattie, and Bresnahan (BBB) scale and the horizontal ladder test) was significantly improved in the RADA16/SP group compared to the control. Our findings indicate that RADA16 with substance P effectively stimulates angiogenesis within the damaged spinal cord and is a possible candidate for post-SCI functional recovery. Funding: This work was supported by the National Research Foundation (NRF) and the Ministry of Science and ICT (MSIT) (2019R1A6A1A11034536, 2020R1A2C2004764); the Korea Medical Device Development Fund grant funded by the Korean government (the Ministry of Science and ICT; the Ministry of Trade, Industry and Energy; the Ministry of Health & Welfare; the Ministry of Food and Drug Safety) (202017D01); the Ministry of Trade, Industry and Energy (MOTIE) (20008686); and the KU-KIST Graduate School of Converging Science and Technology Program, Republic of Korea. Declaration of Interests: The authors declare no conflicts of interest. Ethics Approval Statement: All animal experiments were approved by the Institutional Animal Care and Use Committee of Dankook University (Approval No. 19-010).

BackgroundTumor associated macrophages (TAMs) play a critical role in tumor immunosuppression and resistance to immune checkpoint blockade. Reprogramming ‘M2’ TAMs to a proinflammatory ‘M1’ phenotype by selectively silencing M2 phenotype-driving transcription factors, such as STAT6, is a promising strategy to relieve TAM-induced immunosuppression. We have developed exoASO-STAT6™, an investigational therapeutic candidate consisting of exosomes loaded with antisense oligonucleotides (ASOs) targeting STAT6. By leveraging the TAM tropism of exosomes, exoASO-STAT6™ is the first systemically administered exosome designed to selectively silence STAT6 in TAMs. Preclinical biodistribution studies demonstrated that the liver is the main organ targeted by exoASO after intravenous (IV) dosing.MethodsWe evaluated the translational potential of exoASO-STAT6 to treat hepatocellular carcinoma through pharmacokinetics (PK), pharmacodynamic (PD) and anti-tumoral efficacy studies in preclinical models, as well as computational analysis of human HCC datasets.ResultsPK/PD were evaluated in naïve mice dosed IV with a single dose of exoASO-STAT6. PK analysis showed that the STAT6-ASO is rapidly cleared from plasma. Retention of the ASO in liver was dose-dependent and observed for at least 21 days. exoASO-STAT6 induced significant Stat6 mRNA knockdown (KD) in liver tissue with maximum KD at day 7 (70% KD at the 30 ug dose). IV administration of exoASO-STAT6 in an orthotopic, CPI resistant HCC model attenuated tumor growth and induced complete remission of tumor lesions in 50% of mice, while combination therapy with anti-PD1 antibodies further enhanced anti-tumor activity (75% complete remissions). Gene expression and histological analysis of the liver showed effective remodeling of the tumor microenvironment including a significant increase in interferon and cytotoxic T-cell gene signatures and iNOS expression. PD studies were also performed in cynomolgus monkeys that demonstrated a dose-dependent and durable silencing of STAT6 mRNA (50% and 31% at 1- and 3- weeks post-dose, respectively). STAT6 knockdown correlated with a reduction in STAT6 target genes, IL4R and EGR2, confirming modulation of the STAT6 pathway. Finally, we identified a STAT6 macrophage transcriptional signature and show high expression in human HCC tumors, both in immune cell-rich and TAM-rich/CD8 T-cell low tumors that correlates with worse survival.ConclusionsIn summary, we demonstrate that exoASO-STAT6 has a durable PK/PD profile in the liver of several species and potent antitumoral efficacy in a preclinical model of HCC. Furthermore, we identify an inverse correlation between the STAT6 macrophage signature and survival in human HCC tumors. Altogether our data support the systemic administration of exoASO-STAT6 as a promising therapy for liver cancer.Ethics ApprovalFor MiceMice were maintained and treated at the animal care facility of Codiak Biosciences in accordance with the regulations and guidelines of the Institutional Animal Care and Use Committee (CB2017-001).Animal housing and experimental procedures (mice) were conducted according to the French and European Regulations and the National Research Council Guide for the Care and Use of Laboratory Animals and Institutional Animal Care and Use Committee of Oncodesign (Oncomet) approved by French authorities (CNREEA agreement N° 91).For cynomolgus monkeys:All animals were maintained and treated at the animal care facility of Altasciences in compliance with the Animal Welfare Act and recommendations set forth in the Guide for the Care and Use of Laboratory Animals (National Research Council 2011).

Background: Nephrotic syndrome (NS) is associated with kidney podocyte injury and may occur as part of thyroid autoimmunity such as Graves’ disease. Therefore, the present study was designed to ascertain if and how podocytes respond to and regulate the input of biologically active thyroid hormone (TH), 3,5,3'-triiodothyronine (T3); and also to decipher the pathophysiological role of type 3 deiodinase (D3), a membrane-bound selenoenzyme that inactivates TH, in kidney disease. Methods: To study D3 function in healthy and injured (PAN, puromycin aminonucleoside and LPS, Lipopolysaccharide-mediated) podocytes, immunofluorescence, qPCR and podocyte-specific D3 knockout mouse were used. Surface plasmon resonance (SPR), co-immunoprecipitation and Proximity Ligation Assay (PLA) were used for the interaction studies. Findings: Healthy podocytes expressed D3 as the predominant deiodinase isoform. Upon podocyte injury, levels of DIO3 transcript and D3 protein were dramatically reduced both in vitro and in the LPS mouse model of podocyte damage. D3 was no longer directed to the cell membrane, it accumulated in the Golgi and nucleus instead. Further, depleting D3 from the mouse podocytes resulted in foot process effacement and proteinuria. Treatment of mouse podocytes with T3 phenocopied the absence of D3 and elicited activation of αvβ3 integrin signaling, which led to podocyte injury. We also confirmed presence of an active thyroid stimulating hormone receptor (TSH-R) on mouse podocytes, engagement and activation of which resulted in podocyte injury. Interpretation: The study provided a mechanistic insight into how D3-αvβ3 integrin interaction can minimize T3-dependent integrin activation, illustrating how D3 could act as a renoprotective thyrostat in podocytes. Further, injury caused by binding of TSH-R with TSH-R antibody, as found in patients with Graves’ disease, explained a plausible link between thyroid disorder and NS. Funding: Funding support from American Thyroid Association (ATA) to NJT (ATA-2018-050.R1). Declaration of Interest: Jochen Reiser has patents on novel strategies for kidney therapeutics and stands to gain royalties from their commercialization. He is the co-founder of Walden Biosciences (Cambridge, MA, USA), a biotechnology company in which he has financial interest, including stock. Other authors have nothing to disclose and there are no competing or conflicting interests. Ethical Approval: All animal experiments were carried out according to the NIH’s Guide for the Care and Use of Experimental Animals (National Academies Press, 2011), and approved by the Institutional Animal Care and Use Committee (IACUC) at Rush University (Chicago, Illinois, USA). Human biopsy kidney sections from healthy donors and patients with FSGS, DN and MCD were procured in accordance with guidelines on human research and with approval of the Institutional Review Board of Rush University Medical Center (Chicago, Illinois, USA).

Aim: Hyperleptinemia potentiates the effects of many atherogenic factors, such as inflammation, platelet aggregation, migration, hypertrophy, proliferation of vascular smooth muscle cells, and endothelial cell dysfunction. The present study analysed the effects of long-term hyperleptinemia in an in vivo myocardial ischemia-reperfusion model to demonstrate whether the in vivo deleterious effect also affects cardiac structure and function. Main methods: Rats by were subcutaneously administered leptin for 8 days to estimate the involvement of the JAK/STAT pathway. Data from 58 male Wistar rats were included in the final analysis. Myocardial infarction (MI) was modelled by the 30-minute ligation of the main left coronary artery followed by 120-minute reperfusion. Hemodynamic measurements, electrocardiography monitoring, echocardiography, myocardial infarct size and area at risk, blood biochemical parameters, leptin, IL-6, TNF-alpha, FGF-21, and cardiomyocyte morphology were measured. Statistical analyses were performed using IBM SPSS Statistics v.26. Key findings: Eight-day hyperleptinemia in rats led to increased an blood pressure and heart rate, myocardial hypertrophy, impaired LV function, an increased frequency of ischemic arrhythmias, dyslipidaemia, systemic inflammation, and an increased size of induced myocardial infarction. Significance: The blockade of the JAK/STAT signalling pathway effectively reversed the negative effects of leptin, including increased blood pressure and total cholesterol. Funding Information: The study has been supported by the grant from the Ministry of Science and Higher Education of the Russian Federation (agreement 075-15-2020-800). Declaration of Interests: None to declare. Ethics Approval Statement: This study was conducted in conformance with the policies and procedures detailed by the “Institutional Animal Care and Use Committee” (IACUC). All animal experimental protocols and procedures were reviewed and approved by the Committee for the Control of the Maintenance and Use of Laboratory Animals of the V. A. Almazov National Medical Research Center (date: 06-FEB-2020, application number: 20-01).

Background: It has known that moderate mechanical loading, like that caused by exercise, promotes bone formation. However its underlying mechanisms remain elusive. Methods: Mice were randomly maintained a sedentary lifestyle in cage or subjected to run on a rodent treadmill for 5 weeks. Bone metabolism of mice was evaluated with micro-CT and immunohistochemistry staining. Bioinformatics analysis was performed to identify the candidate chemokine for mechanical-induced bone formation. Flow cytometry and in vivo and in vitro functional assays were conducted to explore the mechanisms of chemokine promoting mesenchymal progenitors recruitment and bone remodeling. Findings: Moderate running dramatically improved trabecular bone remodeling and increased the number of osteoblasts but had no effect on osteoclasts. A dramatic increase in the number of colony forming unit-fibroblast (CFU-F) in endosteal bone marrow and the percentage of CD45- Leptin receptor+ endosteal mesenchymal progenitors were observed. Bioinformatics analysis identified chemokine c-c-motif ligands (CCL2) as a critical candidate induced by mechanical loading. Interestingly, we found that CCL2 was up-regulated mainly in osteoblastic cells in the tibia of mice after moderate running. Further, we found that mechanical loading up-regulates the expression of CCL2 by activating ERK1/2 pathway, thereby stimulating the migration of endosteal mesenchymal progenitors. Interpretation: These results uncover an unknown connection between osteoblasts and endosteal mesenchymal progenitors recruited in the increased bone formation induced by mechanical loading. Funding Statement: This study was supported by the National Natural Science Foundation of China (81830079), Science and Technology Key Planning Project of Guangdong Province (2019B020201013), Natural Science Foundation of Guangdong (2019A1515110818), and President Foundation of Nanfang Hospital, Southern Medical University (2019Z012). Declaration of Interests: The authors declare that there is no conflict of interests. Ethics Approval Statement: All animal protocols were approved by Institutional Animal Care and Use Committee (IACUC) at Nanfang Hospital, Southern Medical University before study and all experiments were performed in accordance with IACUC guidelines.

Background: Osteosarcoma (OS) is the most common malignant bone cancer. Recently, more studies have shown that TPX2, a potential oncogene considered to be an important prognostic biomarker, is widely expressed in various tumors. However, until now, little is known about the impacts of TPX2 on the pathogenesis of OS. Methods: TPX2 expression of in OS cells and tissues was detected by Western blot and immunohistochemical analyses. Various assays were used to analyze OS cell proliferation and metastasis in vitro and in vivo. The effect of TPX2 on the expression of E2F1 and proteins encoded by E2F1-regulated genes was evaluated by Western blotting. Coimmunoprecipitation, immunofluorescence and in vitro ubiquitination assays were employed to explore the molecular mechanisms underlying the effect of TPX2 on stabilizing E2F1 and maintaining E2F1 signaling. Findings TPX2 is overexpressed in OS and is significantly correlated with the malignant progression of OS. TPX2 probably plays a critical role in the proliferation and metastasis of OS cells in vitro. Furthermore, TPX2 regulates E2F1 signaling by affecting the stability of E2F1, thereby affecting the proliferation and metastasis of OS cells. In addition, PSMD14 impacts the degradation and regulates the stability of E2F1. Most importantly, TPX2 regulates the interaction of E2F1 and PSMD14 to influence E2F1 degradation. Interpretation: Collectively, our research not only identifies TPX2 ground as a promising therapeutic target in cancer but also illuminates its underlying molecular mechanism. We showed that TPX2 promotes the malignant progression of OS by regulating E2F1 signaling. Funding: This project was supported by the NSFC (81802985), and a Research Grant from the Shanghai Hospital Development Center (SHDC2020CR2037B). Declaration of 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. Ethical Approval: All animal experiments were performed under the protocols approved by the Shanghai General Hospital Institutional Animal Care and Use Committee. Six-week-old female BABL/c nude mice were obtained from SLRC laboratory Animal (Shanghai, China). The Central Laboratory of Shanghai General Hospital approved this study.

The Institutional Animal Care and Use Committee (IACUC) of Seoul National University (SNU) plays a key role in monitoring and managing the humane use of animals in scientific research. Here, as one of the pioneers of the IACUC in Korea, we reported SNU-IACUC operations and activities including committee establishment and legal formulation, protocol review, and post-approval monitoring of protocols, which the IACUC has undertaken in the last decade. In addition, legal regulations and improvements were also discussed, and encompassed the limited number of committee members and the single IACUC policy in Korea. As of December, 2020, amendments are on the table at the National Assembly. We also emphasized the independent nature of the IACUC in protecting activities, including approval and monitoring animal experiments, and its public role in narrowing the knowledge gap between society and scientists. Thus, the aim of this report is to help society and scientists understand the operations of the SNU-IACUC and its role in animal welfare.

The need for severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) next-generation vaccines has been highlighted by the rise of variants of concern (VoCs) and the long-term threat of emerging coronaviruses. Here, we design and characterize four categories of engineered nanoparticle immunogens that recapitulate the structural and antigenic properties of the prefusion SARS-CoV-2 spike (S), S1, and receptor-binding domain (RBD). These immunogens induce robust S binding, ACE2 inhibition, and authentic and pseudovirus neutralizing antibodies against SARS-CoV-2. A spike-ferritin nanoparticle (SpFN) vaccine elicits neutralizing titers (ID50 > 10,000) following a single immunization, whereas RBD-ferritin nanoparticle (RFN) immunogens elicit similar responses after two immunizations and also show durable and potent neutralization against circulating VoCs. Passive transfer of immunoglobulin G (IgG) purified from SpFN- or RFN-immunized mice protects K18-hACE2 transgenic mice from a lethal SARS-CoV-2 challenge. Furthermore, S-domain nanoparticle immunization elicits ACE2-blocking activity and ID50 neutralizing antibody titers >2,000 against SARS-CoV-1, highlighting the broad response elicited by these immunogens., Graphical abstract, Joyce et al. generate four categories of engineered SARS-CoV-2 ferritin nanoparticle immunogens using structure-based vaccine design that recapitulate the prefusion SARS-CoV-2 spike, S1, and RBD. These immunogens induce robust and protective neutralizing antibody responses against SARS-CoV-2 and elicit potent neutralization against variants of concern and the heterologous SARS-CoV-1.

Cairo University was the first academic institution in Egypt to establish an Institutional Animal Care and Use Committee (IACUC), as mandated by the World Organisation for Animal Health (OIE). Animal-based research should be performed in accordance with international regulations to monitor the humane care and use of the laboratory animals. Until 2018, the formal training of researchers in the appropriate and correct methods of animal handling during sampling and administration, as well as their husbandry demands, was an uncommon practice in Egypt. In 2018, the Egyptian Association for Animal Research Advancement (EAARA) organised the first international course in laboratory animal science (LAS), in collaboration with Utrecht University (The Netherlands) and the Faculty of Science, Cairo University, to raise researchers’ awareness and increase their knowledge of the principles that govern the humane use and care of laboratory animals. A total of 26 researchers from a number of fields (veterinary medicine, dentistry, science, medicine, pharmacy and agriculture) enrolled in the course. In the responses to the post-course questionnaire, 24 (92.3%) participants stated that the principles of animal welfare (Three Rs) were well explained. In addition, 18 (69%) participants found that the course improved their skills in animal sampling and handling. Of the 26 participants, 22 (84.6%) became aware of their responsibility towards their experimental animals and agreed that the different methods of euthanasia were well explained. In conclusion, the general assessment of the course revealed a positive outcome regarding the culture of animal care; the course was repeated a year later, and several participants were enlisted as trainers in this second course.

Melanoma is known to be a heterogeneous tumor, but the impact of this heterogeneity upon therapeutic response is not well understood. In the current study, we used single cell mRNA analysis and our Single Cell HETerogeneity (SinCHET) platform to define the transcriptional diversity of melanoma and its dynamic response to both BRAF inhibitor therapy and treatment holidays. Our analyses showed melanoma cell lines and patient specimens to be composed of >3 co-existent, transcriptionally distinct states. Among these, State #1 had increased expression of cyclin D1, ERBB3, STAT3/5, MITF and I²-catenin and lower expression of c-JUN and RTKs such as Axl and EGFR. State #2 was characterized by higher expression of ERBB3, Axl and the transcription factor c-JUN. State #3 was characterized by high Axl, c-JUN, E2F1, WEE1, c-MET and EGFR expression and lower expression of MITF, ERBB3 and SMAD9. The cell state composition was dynamically regulated in response to BRAF inhibitor therapy, with State #1 declining and States #2 and #3 increasing in response to drug. The percentage of cells in State #1 recovered following a drug holiday, allowing for a successful therapy rechallenge. Other melanomas that lacked State #1 had varying degrees of intrinsic drug resistance and were not amenable to rechallenge. We next leveraged the differences in fitness between the different transcriptional states in the absence and presence of drug to develop a mathematical model that optimized therapy schedules to retain the drug sensitive population. In vivo validation of the mathematical model demonstrated that the personalized adaptive dosing schedules were better at suppressing tumor growth than either continuous or fixed intermittent BRAF inhibitor schedules. Together our studies provide the first preclinical evidence that transcriptional heterogeneity at the single cell level predicts for the initial sensitivity to BRAF inhibitor therapy. We further demonstrate that manipulating transcriptional heterogeneity through personalized adaptive therapy schedules that can delay the time to resistance. Funding Statement: Supported by SPORE grant P50 CA168536, R21 CA198550, R21 CA216756 (to KSMS) K99 CA226679 (to IS), Moffitt Cancer Center PSOC, U54 CA193489 (EK & AA) and the Cancer Center Support Grant P30 CA076292 from the NIH (NCI/NIH). We also acknowledge the support Miami Center for AIDS Research (CFAR) at the University of Miami Miller School of Medicine funded by P30AI073961. Declaration of Interests: We have no competing interest to declare. Ethics Approval Statement: All animal experiments were carried out in compliance with ethical regulations and protocols approved by the University of South Florida Institutional Animal Care and Use Committee.

The purpose of this work was to determine the safety of inhaled hydrogen gas in healthy animals. Female mice were exposed to medical air with or without hydrogen gas (concentration 2.4%) for 72 hours (n = 25 mice/group). Mice underwent a standardized and validated neurobehavioral examination, SmithKline Beecham, Harwell, Imperial College, Royal London Hospital, Phenotype Assessment (SHIRPA) protocol, prior to and following the exposure period. Blood was withdrawn for serologic evaluation and all major organ tissues were evaluated histologically. The average hydrogen concentration within the chamber was 2.27%. Following exposure, there was no significant change in body weight in either group. Similarly, there was no significant change in the total SHIRPA score, although hydrogen-treated mice exhibited significantly lower spontaneous locomotor activity (P < 0.0001) in a subset of the test; all other aspects of the mouse neurologic exam were normal in hydrogen-treated animals. Brain histopathology was also normal in all mice, as was the histology of all other major organs. There were no significant differences in complete blood count, serum chemistry, or arterial blood gases between control and hydrogen-treated mice (P > 0.05 for all). Hydrogen gas did not appear to cause significant adverse effects when administered to healthy mice for 72 hours, with the possible exception of decreased spontaneous locomotor activity. The study was approved by the Institutional Animal Care and Use Committee at Boston Children’s Hospital, USA (approved number 18-01-3536) on January 25, 2018.

Calorie restriction (CR) has been well proved to be a powerful tool to improve metabolic health associated with aging; and many types of CR have been proposed. Intermittent CR has become a trend in recent years due to its better compliance than continuous CR every day. However, there are few studies that directly compare the interventional efficacy of intermittent CR to continuous CR in metabolic disorders such as diabetes. In this study, we analyzed two regimes of intermittent CR with the calorie-matched continuous CR in two diabetic mouse models including db/db and streptozotocin-treated mice. Intermittent CR was carried out by a fasting-mimicking diet (FMD, with 30% calorie intake of the control per day) for 2 days or 5 days (i.e., 2-5 or 5-9 regimes followed by free eating for 5 or 9 days respectively). In the two diabetic mouse models, both intermittent CR and continuous CR significantly reduced fasting blood glucose level and improved insulin sensitivity. However, intermittent CR performed significantly better than continuous CR in improving glycemic control and insulin sensitivity in db/db mice. In addition, intermittent CR improved the glucose homeostasis of the db/db mice without causing loss of body weight. Analyses with the pancreatic islets reveal that intermittent CR profoundly elevated the number of insulin-positive cells in both diabetic mouse models. In conclusion, our study indicated that both intermittent CR and continuous CR can lower fasting blood glucose level in the diabetic mice, while intermittent CR is better than the latter in improving insulin sensitivity. Funding Statement: This study was supported by research grants from the National Natural Science Foundation of China (31630036 to Y.C.), the Ministry of Science and Technology of China (2016YFA0500103 to Y.C.), and the Chinese Academy of Sciences (XDA12010102, QYZDJ-SSW-SMC008, ZDRW-ZS-2016-8, and Y817X11141 to Y.C Declaration of Interests: The authors declare no conflict of interest. Ethics Approval Statement: The animal experiments were conducted in accordance with the guidelines of the Institutional Animal Care and Use Committee of the Institute for Nutritional Sciences, Shanghai Institutes for Biological Sciences (SIBS), Chinese Academy of Sciences (CAS) with an approval number 2010-AN-8.

Although there are numerous Institutional Animal Care and Use Committee (IACUC) protocols aimed at standardizing the oral administration (gavage) of liquid chemicals to laboratory rodents, there is ample variability across protocols, and there are no protocols intended for the gavage of microbial communities to mice or rats. The objective of this study was to conduct a scoping review of publically available IACUC protocols from institutions in the United States, identify protocol criteria and deficiencies, and generate an ‘analytical consensus’ to unify such criteria into two revised protocols: one for chemicals, and another for microbes in mice. Eighteen (n = 18) written institutional protocols from prominent universities, and 26 demonstrations videos from various sources (accounting for >155,000 views) were identified in the World Wide Web. Although written protocols listed up to five criteria for consideration (dosing/volume, pregnancy, animal weight), collectively there was major variability and poor statistical agreement across methods (0% Kappa, p = 0.98). Because protocols also lacked details relevant for the bypassing and survival of live microorganisms in the gastric (antimicrobial) environment, we compiled two ‘analytical (Kappa-based) consensus’ protocols from available and new criteria, for the ‘gavage of chemicals’ and for the ‘gavage of delicate microscopic microbial communities’ to mice. A major difference lies in the volume of administration, (i.e., 20 mL/kg, without restrictions, after 4-6 h of fasting, for microbes), which was graphically illustrated simulating the predicted impact of administering large and small volumes on microbial distribution of simple microscopic communities. In conclusion, publicly available IACUC gavage protocols are highly variable, and do not reflect the need to adjust the dose volumes to ensure the rapid bypassing of the gastric environment which would impact the survival of microbes, especially if composing delicate microscopic communities. An ‘analytical consensus’ of IACUC-approved protocols is herein presented as a unifying baseline protocol for consideration. Impact statement Institutional protocols designed for the oral administration of live microbial communities, either complex or microscopic (microcosmic), to mice do not exist. However, this approach is increasingly employed by investigators focusing on the gut microbiome in experimental research. Herein, we propose two analytically Kappa-based consensus protocols to promote reproducibility and standardization in research practices and describe biologically relevant factors in achieving optimal microbial engraftment of communities in germ-free mice.

Anterior cruciate ligament (ACL) injuries are frequent, as >200,000 injuries occur in the United States alone each year. Owing to the risks for associated meniscus and cartilage damage, ACL injuries are a significant source of both orthopedic care and research. Given the extended recovery course after ACL injury, which often lasts 1-2 years, and is associated with limited participation in sports and activities of daily living for patients, there is a critical need for the evolution of new and improved methods for ACL repair. Subsequently, animal models of ACL reconstruction (ACLR) play a key role in the development and initial trialing of novel ACL interventions. This article provides a clear operative description and associated illustrations for a validated, institutional animal care and use committee, and veterinarian approved and facile model of ACLR to serve researchers investigating ACLR.

Background: Tumor cells benefit from tumor-associated macrophages (TAMs) promoting tumor growth and modulating functions of other cells in tumor microenvironment (TME). However, how tumor cells regulate the property of TAMs during tumor invasion remains to be defined. Methods: Mouse tumor models and cancer patients' samples were analyzed to determine LAMP2a expression in TAMs. In vitro mouse primary macrophages were used to assess LAMP2a-modulated macrophage activation, and to verify LAMP2a's target proteins. The effect of LAMP2a-knockdown on tumor progression and TME maintaining was determined by using mouse tumor models. Findings: Lysosome associated membrane protein type 2A (LAMP2a) is upregulated in TAMs by tumor cells and important for tumor progression. LAMP2a expression in TAMs, but not in tumor cells, is associated with poor prognosis in breast cancer. LAMP2a inactivation induced by either shRNA or CRISPR/Cas9 prevents TAMs activation and tumor growth. LAMP2a degrades PRDX1 (peroxiredoxin 1) and CRTC1 (CREB-regulated transcription coactivator 1) to promote macrophage pro-tumorigenic activation. Interpretation: Our study suggests that tumor cells utilize LAMP2a-PRDX1/CRTC1 axis to modulate TAMs activation and promote tumor growth, reveals the role of LAMP2a in macrophage study and TAM-targeting tumor immunotherapy. Funding Statement: National Key Research and Development Program of China (No. 2016YFA0201402); National Natural Science Foundation of China (No. 81602492). Declaration of Interests: All authors declare that they have no competing interests. Ethics Approval Statement: All animal experiments were approved by the institutional Animal Care and Use Committee. All patient samples concerned studies were approved by West China Hospital IRB committee. All human subjects (except for tissue microarray) were obtained from patients who were receiving treatment in West China Hospital, with all patients fully informed consent.

Background: The drug-resistance and relapse of diffuse large B-cell lymphoma (DLBCL), which are related to mesenchymal stem cells (MSCs), have become increasingly common. However, the underlying mechanisms remain elusive. Methods: CCK 8 assay, colony formation assay, and xenograft mouse model were used to investigate the effects of hBMSCs on DLBCL growth.Immunohistochemistry, qRT-PCR, and ELISA were used to study the expressions of IL-6 and IL-17A. Flow cytometry was used to analyze Th17 cells and Treg cells expressions. Western blot analysis, microarray analysis, and bioinformatics analysis were used to analyze the pathways of IL-6 or IL-17A mediated DLBCL growth. Findings: HBMSCs promoted DLBCL growth by secreting IL-6 in vitro and in vivo and simultaneously upregulating IL-17A in vitro. IL-6 and IL-17A synergistically promoted the growth and drug-resistance of DLBCL cells by protecting them from spontaneous or drug-induced apoptosis in vitro. IL-6 or IL-17A activated the JAK2/STAT3 pathway or upregulated cyclin D2 via activation of PI3K/Akt signaling in vitro, respectively. Interpretation: The present results indicated that hBMSCs might have a "dual effect" on promoting DLBCL progression and drug-resistance by secreting IL-6 and upregulating IL-17A. IL-6, IL-17A, p-STAT3, p-Akt or cyclin D2 may be potential molecular targets for overcoming drug-resistance in patients with relapsed or refractory DLBCL. Funding Statement: This work was supported by the Natural Science Foundation of Guangdong Province, China (2018A0303130167); the Guangzhou Planned Project of Science and Technology, China (201707010279, 201300000100, and 201704020105); the Guangdong Medical Science and Research Foundation, China (A2018258); the Guangzhou General Science and Technology Project of Health and Family Planning, China (20181A011011). Declaration of Interests: The authors declare that they have no competing interests. Ethics Approval Statement: This research was approved by the Ethics Committee of Guangzhou First People’s Hospital (K-2017-066-02). Written informed consent was obtained from all participants or their families. All animal experimental procedures and protocols were approved by the Institutional Animal Care and Use Committee at Guangzhou First People's Hospital, Guangzhou Medical University.

While animal models of controlled cortical impact often display short-term motor dysfunction after injury, histological examinations do not show severe cortical damage. Thus, this model requires further improvement. Mice were subjected to injury at three severities using a Pin-Point™-controlled cortical impact device to establish secondary brain injury mouse models. Twenty-four hours after injury, hematoxylin-eosin staining, Fluoro-Jade B histofluorescence, and immunohistochemistry were performed for brain slices. Compared to the uninjured side, we observed differences of histopathological findings, neuronal degeneration, and glial cell number in the CA2 and CA3 regions of the hippocampus on the injured side. The Morris water maze task and beam-walking test verified long-term (14–28 days) spatial learning/memory and motor balance. To conclude, the histopathological responses were positively correlated with the degree of damage, as were the long-term behavioral manifestations after controlled cortical impact. All animal procedures were approved by the Institutional Animal Care and Use Committee at Shanghai Jiao Tong University School of Medicine.

In fiscal year 2016, agricultural animals such as swine, sheep, goats, and cattle represented 10% of the 820 812 animals used in USDA-regulated research. In addition to traditional agricultural animals, research studies using captive wildlife are becoming increasingly important as human and livestock populations encroach upon, and thus expand interactions with, wildlife populations on the landscape. Optimum healthcare of both livestock and captive wildlife in a research setting requires proper husbandry, management, and veterinary care. Regardless of animal species, proper care and management are essential for animal well-being, valid research data, and the health and safety of animal care personnel. Using wildlife in research presents unique challenges as there is generally limited peer-reviewed research on wildlife welfare, husbandry, and nutrition. Animals often become excited during handling or transport, and care must be taken to avoid injury. When severe injuries do occur, differences may exist in methods of euthanasia. Many wildlife species are evolutionarily programmed to conceal signs of illness, making assessment of their condition difficult; moreover, attending veterinarians are often not as experienced in the care of wildlife as they are in the care of traditional laboratory animals or livestock. These differences are further magnified in the context of wildlife field research. The concepts of replace, reduce, and refine are as valid in livestock and wildlife research as in biomedical research, and investigators should work closely with their Institutional Animal Care and Use Committees to ensure humane animal care. The Institutional Animal Care and Use Committee is centrally important in providing guidelines relative to ethical use of animal subjects for research and can serve as a valuable resource for research accountability.

There have been recent efforts to reduce the administrative burden imposed on investigators. Although a complete and thorough review of proposed animal studies is an essential function of the Institutional Animal Care and Use Committee (IACUC), efforts to streamline and clarify this process may help investigators spend less time writing animal use protocols and responding to committee comments. The IACUC relies on well-written protocols for an efficient review process. A well-designed protocol form is also critical in guiding investigators through the process. However, it is ultimately the investigators’ responsibility to ensure that the information they provide answers all the IACUC’s questions with enough detail and quality for a fast and effective review. This article, aimed primarily for researchers but also IACUC administrators, provides an overview of the IACUC protocol review and approval process, the criteria that the IACUC uses for evaluations, and the type of information that should be included in the various sections of the protocol form. Some specific examples are also provided.

BackgroundProgrammed death-ligand 1 (PD-L1) and programmed cell death protein 1(PD-1) blockade therapy has revolutionized the treatment landscape of malignancies. However, only a minority of patients are anticipated to experience a deep and durable response. In addition, successful therapeutic agonism of 4-1BB, a promising co-stimulatory immunologic target, has been limited by major safety concerns of hepatotoxicity or suboptimal agonistic potency. ATG-101, a novel PD-L1/4-1BB bispecific antibody, was designed to activate 4-1BB positive T cells in a PDL1-crosslinking dependent manner and to effectively treat tumors without on-target-off-tumor liver toxicity (figure 1).MethodsATG-101 was developed by introducing lower affinity 4-1BB scFv into a human IgG1 PD-L1 monoclonal antibody. The N297A mutation on CH2 abolishes the binding capacity to most FcγRs but retains the binding to FcγRn. A series of in vitro and in vivo studies were performed to evaluate the potency, safety and specific mechanism of action.ResultsATG-101 simultaneous binds to 4-1BB and PD-L1 with higher affinity to PD-L1, and potently activates 4-1BB positive T cells when crosslinked by PD-L1 positive cells. Upon crosslinking, ATG-101 also activates PD1+TIM3+ exhausted T cells in vitro, suggesting a potential in reversing T-cell dysfunction and exhaustion (figure 1). ATG-101 shows potent anti-tumor activities in various animal models, including h4-1BB humanized mice bearing MC38 colon cancer, PD(L)1 blockade insensitive B16F10 melanoma and EL4 lymphoma, with no body weight loss observed. To evaluate ATG-101 efficacy in tumors progressing after anti-PD(L)1 treatment, mice bearing MC38 tumors were treated with anti-PDL1 initially to achieve tumor growth inhibition, and half of the mice switched to ATG-101 upon disease progression, the other mice continuing with anti-PD-L1 treatment. ATG-101 induced potent tumor growth inhibition and tumor regression in anti-PDL1-resistant tumors and prolonged survival. Flow cytometry and multiplex IHC staining of tumor samples from mice treated with ATG-101 or control suggest that ATG-101 increases the infiltration, proliferation and activation of CD8+ T cells (figure 2), the infiltration of natural killer T cells and the CD8+/Treg ratio in TILs. In a 4-week GLP toxicity study in cynomolgus monkey, up to 100mg/kg repeated doses of ATG-101 were well tolerated with no hepatotoxicity observed.Abstract 893 Figure 1ATG-101 conditionally activates exhausted T cells. (A) Mechanism of action of ATG-101 (B) Exhausted T cells were induced by CD3+T cells cultured with anti-CD3/CD28 beads for 6 days. The percentage of terminally exhausted T cells (PD-1+Tim-3+) and progenitor exhausted T cells (PD-1+Tim-3-) were increased on Day6 (C) With the presence of PD-L1 positive cells, ATG-101 induced the IL2 and INF-γ secretion by exhausted T cells.Abstract 893 Figure 2Potent in vivo efficacy of ATG-101. (A) Representative MC38 tumor growth curve for individual mice treated with PBS (black), 10mpk atezolizumab (Atezo) only (red) or mice initially treated with 10mpk atezolizumab and switched to 13mpk ATG-101 upon disease progression (red-green); the arrow indicates the day switching Atezo to ATG-101; (B) MC38 tumor growth curve for all individual mice treated with PBS (black, n=6), atezolizumab only (red, n=14), and atezolizumab initially before switching to ATG-101 upon disease progression (red-green, n=14) ; (C) Survival data of mouse shown in (B); (D) Representative images for multiplex IHC staining of tumor samples collected from mouse from (B). (E) Quantitative analysis of TILs shown in (D). Compared with PBS group or atezo-only group, ATG-101 significantly increased the infiltration of T cells in the tumor microenvironment. MHCII: APC Cells, CD4+ CD8-: Helper T Cells, CD4- CD8+: Cytotoxic T Cells, CD4- CD8- F4/80- PDL1+: Tumor Cells.ConclusionsATG-101 demonstrated significant anti-tumor activity in various tumor models including those progressing on anti PD(L)1 treatment. Good safety and PK/PD properties has been demonstrated in preclinical in vivo models. A phase I, multicenter, dose-escalating clinical trial evaluating ATG-101 in patients with solid tumors and hematologic malignancies is ongoing.Ethics ApprovalThe protocol and any amendment(s) or procedures involving the care and use of animals in this study were reviewed and approved by the Institutional Animal Care and Use Committee (IACUC) of CrownBio or Innostar prior to execution with an AUP number or IACUC approval number for each animal study. During the study, the care and use of animals were conducted in accordance with the regulations of the Association for Assessment and Accreditation of Laboratory Animal Care (AAALAC).All studies were conducted following an approved IACUC protocol. AUP NO.:2004-12-1465, 2004-12-1000; IACUC approval number: IACUC-2021-M-003

The epigenetic treatment by 3-Deazaneplanocin A (DZNep), a histone methyltransferase inhibitor, shows great potential to treat acute myeloid leukemia (AML). However, the variant sensitivity and incomplete response to DZNep are very common in different AML cell lines as well as primary leukemia cells from AML patients. Here, we reveal that vitamin C (Vc) dramatically promotes DZNep response in leukemia cells from different AML cell lines and primary AML patient. Vc enhances apoptosis and differentiation induced by DZNep in different AML cell lines in vitro and reduces leukemia progression in vivo. At the molecular level, Vc promotes downregulation of D-3-phosphoglycerate dehydrogenase (PHGDH), an enzyme for serine synthesis, and other anti-apoptotic genes, such as BCL2. Over-expression of PHGDH reverses the enhancing effect of Vc on the anti-leukemia effect of DZNep in AML cells. Together, our findings provide an effective approach to reduce the epigenetic treatment resistance by Vc and would be valuable in potential applications. Funding Information: This work was supported by the National Natural Science Foundation of China (81700149), Guangdong Basic and Applied Basic Research Foundation (No. 2020A1515010199), and the Fundamental Research Funds for the Central Universities (No. 20ykpy30). Declaration of Interests: The authors declare no competing financial interests. Ethics Approval Statement: All samples were collected after obtaining informed consent in accordance with the Declaration of Helsinki. The study was approved by the ethics committees of the Institutional Review Boards. All animal experiments were performed according to protocols approved by the institutional animal care and use committee.

In this essay, I discuss my experience as a member of my university’s Institutional Animal Care and Use Committee (IACUC). Using Blum’s (2016) model of experiential learning (or “learning in the wi...

Precise assessment of spinal cord cystic lesions is crucial to formulate effective therapeutic strategies, yet histological assessment of the lesion remains the primary method despite numerous studies showing inconsistent results regarding estimation of lesion size via histology. On the other hand, despite numerous advances in micro-computed tomography (micro-CT) imaging and analysis that have allowed precise measurements of lesion size, there is not enough published data on its application to estimate intraspinal lesion size in laboratory animal models. This work attempts to show that micro-CT can be valuable for spinal cord injury research by demonstrating accurate estimation of syrinx size and compares between micro-CT and traditional histological analysis. We used a post-traumatic syringomyelia rat model to compare micro-CT analysis to conventional histological analysis. The study showed that micro-CT can detect lesions within the spinal cord very similar to histology. Importantly, micro-CT appears to provide more accurate estimates of the lesions with more measures (e.g., surface area), can detect compounds within the cord, and can be done with the tissue of interest (spinal cord) intact. In summary, the experimental work presented here provides one of the first investigations of the use of micro-CT for estimating the size of intraparenchymal cysts and detecting materials within the spinal cord. All animal procedures were approved by the University of Akron Institutional Animal Care and Use Committee (IACUC) (protocol # LRE 16-05-09 approved on May 14, 2016).

COVID-19 has proven to be a metabolic disease resulting in adverse outcomes in individuals with diabetes or obesity. Patients infected with SARS-CoV-2 and hyperglycemia suffer from longer hospital stays, higher risk of developing acute respiratory distress syndrome (ARDS), and increased mortality compared to those who do not develop hyperglycemia. We find the prevalence of hyperglycemia in patients with COVID-19 ARDS to be equal to those with non-COVID-19 ARDS. Nevertheless, the pathophysiological mechanism(s) of hyperglycemia in COVID-19 remains poorly characterized. Here we show that insulin resistance rather than pancreatic beta cell failure is the prevalent cause of hyperglycemia in COVID-19 patients with ARDS, independent of glucocorticoid treatment. A screen of protein hormones that regulate glucose homeostasis reveals that the insulin sensitizing adipokine adiponectin is reduced in hyperglycemic COVID-19 patients. Hamsters infected with SARS-CoV-2 also have diminished expression of adiponectin. Together these data suggest that adipose tissue dysfunction may be a driver of insulin resistance and adverse outcomes in acute COVID-19. Funding Information: This work was supported by the following grants: NIH R01 DK121140 (J.C.L.) and R01 DK121844 (J.C.L.), NCI R01 CA234614 (R.E.S), NIAID 2R01 AI107301 (R.E.S), and NIDDK R01 DK121072 (R.E.S). R.E.S. and J.C.L are supported as Irma Hirschl Trust Research Award Scholars. In addition, this study received support from NewYork-Presbyterian Hospital (NYPH) and Weill Cornell Medical College (WCMC), including the Clinical and Translational Science Center (CTSC) (UL1 TR 0002384). Declaration of Interests: R.E.S. is on the scientific advisory board of Miromatrix Inc. R.E.S. is a speaker and consultant for Alnylam Inc. The other authors have no conflict of interest. Ethics Approval Statement: The registry was approved by the institutional review board of WCMC (1405015116, 20-05022072, 20-03021681). All animal experiment procedures, breeding, and ethical use were performed in accordance with the guidelines set by the Institutional Animal Care and Use Committee at Mount Sinai School of Medicine.

With the recent upswing of infectious disease outbreaks (coronavirus, influenza, Ebola, etc), there is an ever-increasing need for biocontainment animal use protocols to better address the research of emerging diseases and to increase the health of both animals and humans. It is imperative that we as a research community ensure these protocols are conducted with the utmost scrutiny and regulatory compliance for the welfare of the animals as well as the health and safety concerns of the individual conducting these studies. Both the welfare of the animals and the health and safety of the research staff must be balanced with the integrity of the science being studied. Even prior to reviewing biocontainment protocols, the research stakeholders should have professional and collegial interactions across all levels of the proposed project. These stakeholders should include the attending veterinarian, the principal investigator, the sponsor, and any organic institutional health and safety assets (environmental health and safety, occupational health, biosafety personnel, medical personnel, facilities operations and maintenance, etc). At most institutions, these stakeholders are members of the Institutional Animal Care and Use Committee and may not possess the necessary tools to properly assess an Animal Biosafety Level 3 and 4 animal use protocol. It is the goal of this article to review some basic concepts of biocontainment, discuss critical communications and preapprovals, clinical observations, medical interventions and supportive care, scientific and study endpoints, euthanasia criteria, animal manipulations, documentation, training, emergency response and contingency plans, security, and decontamination and provide a scenario-based and informative thought-provoking process Institutional Animal Care and Use Committee members and veterinary staff may consider during Animal Biosafety Level 3 and 4 protocol review. These topics will enhance the ability of all stakeholders to balance the protection of the people with the integrity of the science and ultimately the welfare of the animal.

Recently approved vaccines have shown remarkable protection in limiting SARS-CoV-2 associated disease. However, immunologic mechanism(s) of protection, and how boosting alters immunity to wildtype and newly emerging strains, remains incompletely understood. Here we profiled the humoral immune response in a cohort of non-human primates immunized with a recombinant SARS-CoV-2 spike (S) glycoprotein (NVX-CoV2373) at two dose levels, administered as a one or two-dose regimen with a saponin-based adjuvant Matrix-M™. While antigen dose had minimal effects, boosting significantly altered the humoral response, driving unique vaccine-induced antibody fingerprints. Differences in antibody effector functions and neutralization were associated with protection in the upper and lower respiratory tract, pointing to compartment-specific determinants of protective immunity against infection. Moreover, NVX-CoV2373 elicited antibodies targeting emerging SARS-CoV-2 variants. Collectively, the data presented here suggest that a single dose may prevent disease, but that two doses may be essential to block further transmission of SARS-CoV-2 and emerging variants. Funding: This work was funded by Operation Warp Speed. We thank Colin Mann and Kathryn Hastie for production of Spike antigens. We thank Nancy Zimmerman, Mark and Lisa Schwartz, an anonymous donor (financial support), Terry and Susan Ragon, and the SAMANA Kay MGH Research Scholars award for their support. We acknowledge support from the Ragon Institute of MGH, MIT and Harvard, the Massachusetts Consortium on Pathogen Readiness (Mass CPR), the NIH (3R37AI080289-11S1, R01AI146785, U19AI42790-01, U19AI135995-02, U19AI42790-01, 1U01CA260476 – 01, CIVIC75N93019C00052), National Science Foundation Graduate Research Fellowship Grant No. #1745302, the Gates foundation Global Health Vaccine Accelerator Platform funding (OPP1146996 and INV-001650), and the Musk Foundation. Conflict of Interest: NP, MGX, JHT, BZ, SM, AMG, MJM, ADP, GG, GS, and LE are current or past employees of Novavax, Inc. and have stock options in the company. GA is the founder of Serom Yx Systems, Inc. AZ is a current employee of Moderna, Inc. but conducted this work before employment.Any opinion, findings, and conclusions or recommendations expressed in this material are those of the author(s) and do not necessarily reflect the views of the National Science Foundation. YG, RC, MJG, CA, KMP, CL, DY, KB, MEM, JL, DM, CM, SS, FA, FK, EOS, DL, and MBF declare no competing interest. Ethical Approval: The work was conducted in accordance with a protocol approved by Texas Biomed’s Institutional Animal Care and Use Committee. All subjects signed informed consent and safety oversight was monitored by a data monitoring board.

Background: Lumbar intervertebral disc (IVD) herniation causes severe low back pain (LBP), which results in substantial financial and emotional strains. Despite the effectiveness of discectomy, there is no existing treatment for post-operative LBP induced by progressive IVD degeneration. Two key factors of LBP are intradiscal inflammation, indicated by tumour necrosis factor alpha (TNF-α) and interleukin-6 (IL-6), and sensory nerve ingrowth into the inner layer of the annulus fibrosus, triggered by nerve growth factor/high-affinity tyrosine kinase A (TrkA) signalling. In an animal models of discectomy, the bioresorbable ultra-purified alginate (UPAL) gel with an extremely low-toxicity has been effective in acellular tissue repair. Methods: We aimed to investigate whether UPAL gel can alleviate LBP using a rat nucleus pulposus (NP) punch model and a rabbit NP aspirate model. In both models, we assessed TNF-α and IL-6 production and TrkA expression within the IVD by immunohistochemistry. Further, histological analysis and behavioural nociception assay were conducted in the rat model. Findings: UPAL gel implantation suppressed TNF-α and IL-6 production, downregulated TrkA expression, inhibited IVD degeneration, and reduced nociceptive behaviour. Interpretation: Our results suggest the potential of UPAL gel implantation as an innovative treatment for IVD herniation by reducing LBP and preventing IVD degeneration after discectomy. Funding Statement: Ministry of Education, Culture, Sports, Science, and Technology of Japan, Akiyama Life Science Foundation, Mochida Pharmaceutical Co., Ltd., and Takeda Science Foundation. Declaration of Interests: The authors declare that no competing interests exist. Ethics Approval Statement: All animal procedures were performed in accordance with the ARRIVE guideline and National Institutes of Health guide for the care and use of Laboratory animals, and the study was approved by the Institutional Animal Care and Use Committee of Hokkaido University (17-0122).

Several SARS-CoV-2 lineages have emerged leading to the divergence of more transmissible variants termed Variants of Concern (VOCs). The natural selection of mutations in the spike protein can impact viral cell entry, transmission, and pathogenesis. Here, we characterized emerging SARS-CoV-2 spike polymorphisms in vitro and in vivo to demonstrate that the substitution S:655Y, included in the highly prevalent Gamma variant, enhances viral replication and spike protein cleavage. Moreover, viral competition experiments demonstrate that the S:655Y transmits more efficiently than the ancestor 655H in the hamster model. Finally, we analyze a set of emerging SARS-CoV-2 variants to investigate how different sets of mutations may impact spike processing. All VOCs tested exhibit an increased spike cleavage and fusogenic capacity. This study demonstrates that the S:655Y is an important adaptative mutation that increases viral cell entry, transmission, and host susceptibility. Moreover, SARS-CoV-2 VOCs show a convergent evolution that promotes the spike protein processing. Funding Information: This research was partly funded by CRIPT (Center for Research on Influenza Pathogenesis and Transmission), a NIAID funded Center of Excellence for Influenza Research and Response (CEIRR, contract #75N93021C00014) (AGS), NCI SeroNet grant U54CA260560 (AGS), NIAID grants U19AI135972 and U19AI142733 (AGS), DARPA grant HR0011-19-2-0020 (AGS), JPB Foundation (AGS), Open Philanthropy Project (research grant 2020-215611 (5384) (AGS), anonymous donors to AGS, NBAF Transition Funds from the State of Kansas (JAR), NIAID Centers of Excellence for Influenza Research and Surveillance under contract number HHSN 272201400006C (JAR), AMP Core of the Center for Emerging and Zoonotic Infectious Diseases of the National Institute of General Medical Sciences (NIGMS) of the National Institutes of Health under award number P20GM130448 (JAR) and Department of Homeland Security Center of Excellence for Emerging and Zoonotic Animal Diseases under grant number HSHQDC 16-A-B0006 (JAR). AGR is funded by Marion Alban MSCIC Scholars Award and the 2020 Robin Chemers Neustein Postdoctoral fellowship. ML is funded by a fellowship of the Belgian American Education Foundation Declaration of Interests: The A.G.-S. laboratory has received research support from Pfizer, Senhwa Biosciences, Kenall Manufacturing, Avimex, Johnson & Johnson, Dynavax, 7Hills Pharma, N-fold LLC, Pharmamar, ImmunityBio, Accurius, Nanocomposix, Hexamer and Merck, outside of the reported work. A.G.-S. has consulting agreements for the following companies involving cash and/or stock: Vivaldi Biosciences, Contrafect, 7Hills Pharma, Avimex, Vaxalto, Pagoda, Accurius, Esperovax, Farmak, Applied Biological Laboratories and Pfizer, outside of the reported work. A.G.-S. is inventor on patents and patent applications on the use of antivirals and vaccines for the treatment and prevention of virus infections, owned by the Icahn School of Medicine at Mount Sinai, New York. The Icahn School of Medicine at Mount Sinai has filed a patent application relating to SARS-CoV-2 serological assays, which lists Viviana Simon as co-inventor. Mount Sinai has spun out a company, Kantaro, to market serological tests for SARS-CoV-2. All other authors have nothing to declare. Ethics Approval Statement: Human SARS-CoV-2: Nasopharyngeal swab specimens were collected as part of the routine SARS-CoV-2 surveillance conducted by the Mount Sinai Pathogen Surveillance program (IRB approved, HS#13-00981). All hamster animal studies were approved by the Institutional Animal Care and Use Committee (IACUC) of Icahn School of Medicine at Mount Sinai (ISMMS). The Institutional Animal Care and Use Committee (IACUC) of the Icahn School of Medicine at Mount Sinai (ISMMS) reviewed and approved the mink model of COVID-19.

The biosafety level-3 (BSL-3) requirement to culture severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is a bottleneck for research and countermeasure development. Here we report a trans-complementation system that produces single-round infectious SARS-CoV-2 that recapitulates authentic viral replication. We demonstrate that the single-round infectious SARS-CoV-2 can be used at BSL-2 laboratories for high-throughput neutralization and antiviral testing. The trans-complementation system consists of two components: a genomic viral RNA containing a deletion of ORF3 and envelope gene, and a producer cell line expressing the two deleted genes. Trans-complementation of the two components generates virions that can infect naive cells for only one round, but does not produce wild-type SARS-CoV-2. Hamsters and K18-hACE2 transgenic mice inoculated with the complementation-derived virions exhibited no detectable disease, even after intracranial inoculation with the highest possible dose. The results suggest that the trans-complementation platform can be safely used at BSL-2 laboratories for research and countermeasure development. Funding: P.-Y.S. was supported by NIH grants AI142759, AI134907, AI145617, and UL1TR001439; awards from the Sealy & Smith Foundation, Kleberg Foundation, John S. Dunn Foundation, Amon G. Carter Foundation, Gilson Longenbaugh Foundation, and Summerfield Robert Foundation; and fund in sponsored research agreement from Q2 Solutions. M.S.D. was supported by R01 AI157155. V.D.M. was supported by NIH grants U19AI100625, R00AG049092, R24AI120942, and a STARs Award from the University of Texas System. S.C.W. was supported by NIH grant R24 AI120942. J.L. is supported by the postdoctoral fellowship from the McLaughlin Fellowship Endowment at UTMB. P.R. and X.X. were partially supported by the Sealy & Smith Foundation. Conflict of Interest: X.Z., X.X., and P.-Y.S. have filed a patent on the trans-complementation system of SARS CoV-2. M.S.D. is a consultant for Inbios, Vir Biotechnology, NGM Biopharmaceuticals, and Carnival Corporation, and on the Scientific Advisory Boards of Moderna and Immunome. The Diamond laboratory has received unrelated funding support in sponsored research agreements from Moderna, Vir Biotechnology, and Emergent BioSolutions. Ethical Approval: Animal studies were carried out in accordance with the recommendations in the Guide for the Care and Use of Laboratory Animals of the National Institutes of Health. The protocols were approved by the Institutional Animal Care and Use Committee at the Washington University School of Medicine (assurance number A3381–01). The research protocol for use of human serum specimens was approved by the University of Texas Medical Branch (UTMB) Institutional Review Board (IRB protocol number 20-0070). All human serum specimens were obtained at the UTMB with patient information de-identified. All aspects of this study were approved by the Institutional Biosafety Committee of the University of Texas Medical Branch at Galveston before the initiation of this study.

Clinical antibiotic-based therapy for H. pylori infections can induce bacterial drug resistance. How to eradicate H. Pylori in vivo exists great challenges. Herein we report that monoclonal antibody conjugated liposomes filled with ICG(HpAb-LiP-ICG) was successfully developed and used for targeted photoacoustic imaging directed sonodynamic therapy (SDT) of H. pylori in vivo. After prepared HpAb-LiP-ICG was orally administered to H. pylori infected mice, photoacoustic imaging clearly showed prepared ICG liposomes could recognize and target actively H. Pylori in gastric organ in mice models, then under ultrasonic radiation on the stomach of mice in vitro, ICG in HpAb-LiP-ICG combined on the surface of H. pylori was activated to produce enough singlet oxygen ( 1O2 )to kill H. Pylori. Pathological analysis showed that H. pylori were killed under antibacterial effect of SDT, gastric mucous tissues recovered to normal status after a month of SDT. In conclusion, prepared HpAb-LiP-ICG nanoprobes-based SDT provides a novel strategy to treat H. pylori infection in vivo, own clinical translational prospects in near future. Funding Information: This work was supported from National Key Research and Development Program of China (2017FYA0205304), National Natural Science Foundation of China (No. 81225010, 81028009, 81921002, 8202010801, 31170961), Shanghai Jiao Tong University Medicine-Engineering Cross budget (No. YG2016ZD10), Shanghai Engineering Research Center for Intelligent diagnosis and treatment instrument (No.15DZ2252000). Declaration of Interests: The authors declare that they have no competing interest. Ethics Approval Statement: All animal experiments were approved by the Institutional Animal Care and Use Committee of Shanghai Jiao Tong University.

Mutations in the SARS-CoV-2 spike receptor-binding domain (RBD) may alter the viral host tropism and affect the activities of neutralizing antibodies (nAbs). Here, we investigated 153 RBD mutants and 10 globally circulating VOC/VOI variants for their cross-species infection potentials and sensitivities to nAbs. Several RBD mutations strengthened viral infectivity in cells-expressing ACE2 orthologs of non-human animals, particularly those less susceptible to the ancestral strain. Antigenic profiles revealed the mutations surrounding aa 439-448 and aa 484 were more likely to cause neutralization resistance. Strikingly, the enhanced cross-species infection potentials to the mouse and ferret, instead of the neutralization-escape scores of the mutations, accounted for the positive correlation with the current cumulative prevalence of mutations in humans. Our results provided prospective landscapes for the cross-species transmission potentials and antigenic drift of circulating spike mutants and highlighted the possible spillover of variants to originally-insusceptible animals in associating with their quick spread worldwide. Funding: This study was supported by the National Natural Science Foundation of China 81902057 (to Y.L.Z.), U1905205 (to Q.Y.), and 81871316 (to Q.Y.) and the CAMS Innovation Fund for Medical Sciences (No. 2019RU022 to N.X.). Declaration of Interests: None to declare. Ethics Approval Statement: The study was approved (number: SPHIRB-202101) by the institutional review board of the School of Public Health of Xiamen University following the Declaration of Helsinki, and written informed consent was obtained.Animal studies were carried out in strict accordance with the recommendations of the Guide for the Care and Use of Laboratory Animals, and were conducted under the approval of the Institutional Animal Care and Use Committee of Xiamen University (mouse experiments) or JOINN Laboratories, Inc. (monkey experiments).

Major depressive disorder (MDD) is a neurasthenic disease, which is the second-largest burden of disease globally. Increasing studies have revealed that depression is associated with abnormalities in gut microbiota and metabolites. Several species of bacteria have been classified as psychobiotics, which confer mental health benefits through interactions with commensal gut microbiota. Therefore, it is essential to identify new psychobiotics and elucidate their mechanisms in the the treatment of depression. This study aims to evaluate the antidepressant effect of Akkermansia muciniphila (AKK) in a mouse model of depression induced by chronic restraint stress (CRS). C57BL/6 male mice were divided into three groups: mice subjected to CRS, mice not subjected to CRS, and mice treated with AKK for three weeks. Behavioral tests were performed, and hormone, neurotransmitter, and brain-derived neurotrophic factor (BDNF) levels were measured. Cecal microbiota was analyzed using 16S rRNA gene sequencing, and serum metabolites were detected using untargeted metabolomics. In addition, correlations between altered gut microbiota and metabolites with significant variations in serum associated with AKK ameliorating depression were analyzed using Pearson’s Correlation coefficient. The results revealed that AKK significantly ameliorated depressive-like behavior and restored abnormal variations in depression related molecular (corticosterone, dopamine and BDNF). Moreover, AKK altered chronic stress-induced gut microbial abnormalities. Untargeted metabolomics analysis revealed 23 potential biomarkers in serum that could be associated with the mechanisms underlying CRS-induced depression and the therapeutic effects of AKK. Pearson's correlation coefficient analysis revealed that AKK predominantly upregulated β-Alanyl-3-methyl-L-histidine and edaravone to relieve depression. Furthermore, β-Alanyl-3-methyl-L-histidine and edaravone exhibited the antidepressant phenotype in mice subjected to CRS. In conclusion, the study demonstrated that AKK ameliorates chronic stress-induced depressive symptoms in mice by regulating gut microbiota and metabolites. Funding Statement: This work was supported by the National Natural Science Foundation of China (No. 81873309). Declaration of Interests: There are no conflicts of interest to declare. Ethics Approval Statement: The experiments were conducted in accordance with the Guide for the Care and Use of Laboratory Animals and were approved by the Institutional Animal Care and Use Committee of the Nanjing University of Chinese Medicine.

Cytokine release syndrome (CRS) is a severe complication of infectious diseases such as coronavirus disease 2019 (COVID-19) that cause serious damage to public health. Currently, no effective therapeutic strategy exists for CRS treatment in clinic, and supportive therapy is still the main method. Here, we show the potential of macrophage membrane-biomimetic nanoparticles for CRS treatment. By fusing macrophage membrane on the surface of the PLGA skeleton, we constructed biomimetic nanoparticles that inherited the membrane receptors from the "parental" macrophages, enabling the neutralization of CRS-related cytokines. We compared three types of macrophage membranes to screen out more effective biomimetic nanoparticles for CRS treatment. Our results show that biomimetic nanoparticles containing M0 macrophage membrane can decrease the levels of pro-inflammatory cytokines involved in CRS to the greatest extent. In a mouse model of RNA virus analog-induced CRS, the macrophage membrane-biomimetic nanoparticles manifested significant therapeutic efficacy by reducing cytokine levels and alleviating organ damage. Funding Information: This work was supported by Scientific Research Project of Tongji University (22120200021), the National Natural Science Foundation of China (82073385 and 81803078), Shanghai Science and Technology Committee (20DZ2255200 and 21140901900). Declaration of Interests: The authors declare no conflict of interest. Ethics Approval Statement: All animal studies were approved under the guidelines of Tongji University Institutional Animal Care and Use Committee.

SARS-CoV-2 frequently causes systemic infection by breaking the bounds of the respiratory systems. Although the development of animal models recapitulating the clinical manifestations of systemic COVID-19 is of utmost importance to understand its pathogenesis, there has not been developed an animal model for systemic infection of SARS-CoV-2. Here we report that Phodopus roborovskii SH101, a laboratory inbred hamster strain of P. roborovskii, infected with SARS-CoV-2 breached the bounds of the respiratory system infection causing systemic infection as in the case of the human counterpart. P. roborovskii SH101 post-infection of SARS-CoV-2 represented most clinical symptoms of COVID-19 such as snuffling, dyspnea, cough, labored breathing, hunched posture, progressive weight loss, and ruffled fur, in addition to high fever accompanied by shaking chills. Histological examinations also revealed a serious right-predominated pneumonia as well as slight organ damages in the brain and liver as in the case of human systemic COVID-19 cases. Funding Statement: This research was supported by JINIS BDRD Research Institute of JINIS Biopharmaceuticals Inc. (Korea). Declaration of Interests: Authors declare no competing interests. Ethics Approval Statement: All procedures involving the mice and hamsters were accredited with the approval of the Institutional Animal Care and Use Committee (IACUC), in compliance with the guidelines of the Ethics Committee of Jeonbuk National University Laboratory Animal Center Guidelines on the Care and Use of Animals for Scientific Purposes. All animal experiments in this study were in accordance with the ARRIVE guidelines and checklist. All the animals used in this research were handled in a manner consistent with CDC/ABSA/WHO guidelines for the prevention of human infection with the SARS-CoV-2 virus.