Generation of Novel Human Red Blood Cell-Bearing Humanized Mouse Models Based on C3-Deficient NOG Mice.
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ABSTRACT:
SUBMITTER: Yamaguchi T
PROVIDER: S-EPMC8353390 | biostudies-literature |
REPOSITORIES: biostudies-literature
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Project description:BackgroundC3 glomerulopathy (C3G) is characterized by the alternative-pathway (AP) hyperactivation induced by nephritic factors or complement gene mutations. Mice deficient in complement factor H (CFH) are a classic C3G model, with kidney disease that requires several months to progress to renal failure. Novel C3G models can further contribute to understanding the mechanism behind this disease and developing therapeutic approaches.MethodsA novel, rapidly progressing, severe, murine model of C3G was developed by replacing the mouse C3 gene with the human C3 homolog using VelociGene technology. Functional, histologic, molecular, and pharmacologic assays characterize the presentation of renal disease and enable useful pharmacologic interventions in the humanized C3 (C3hu/hu) mice.ResultsThe C3hu/hu mice exhibit increased morbidity early in life and die by about 5-6 months of age. The C3hu/hu mice display elevated biomarkers of kidney dysfunction, glomerulosclerosis, C3/C5b-9 deposition, and reduced circulating C3 compared with wild-type mice. Administration of a C5-blocking mAb improved survival rate and offered functional and histopathologic benefits. Blockade of AP activation by anti-C3b or CFB mAbs also extended survival and preserved kidney function.ConclusionsThe C3hu/hu mice are a useful model for C3G because they share many pathologic features consistent with the human disease. The C3G phenotype in C3hu/hu mice may originate from a dysregulated interaction of human C3 protein with multiple mouse complement proteins, leading to unregulated C3 activation via AP. The accelerated disease course in C3hu/hu mice may further enable preclinical studies to assess and validate new therapeutics for C3G.
| S-EPMC7894673 | biostudies-literature
Project description:We developed a novel, rapidly-progressing, severe murine model of C3G by replacing the mouse C3 gene with the human C3 homologue using Velocigene® technology. We conducted functional, histological, molecular and pharmacologic assays to characterize the presentation of renal disease and useful pharmacologic interventions in the humanized C3 (C3hu/hu) mice.
2020-05-20 | GSE150838 | GEO
Project description:C-type lectin-like receptor 2 (CLEC-2) is considered as a potential drug target in settings of wound healing, inflammation, and infection. A potential barrier to this is evidence that CLEC-2 and its ligand podoplanin play a critical role in preventing lymphatic vessel blood filling in mice throughout life. In this study, this aspect of CLEC-2/podoplanin function is investigated in more detail using new and established mouse models of CLEC-2 and podoplanin deficiency, and models of acute and chronic vascular remodeling. We report that CLEC-2 expression on platelets is not required to maintain a barrier between the blood and lymphatic systems in unchallenged mice, post-development. However, under certain conditions of chronic vascular remodeling, such as during tumorigenesis, deficiency in CLEC-2 can lead to lymphatic vessel blood filling. These data provide a new understanding of the function of CLEC-2 in adult mice and confirm the essential nature of CLEC-2-driven platelet activation in vascular developmental programs. This work expands our understanding of how lymphatic blood filling is prevented by CLEC-2-dependent platelet function and provides a context for the development of safe targeting strategies for CLEC-2 and podoplanin.
| S-EPMC8443399 | biostudies-literature
Project description:We have developed NOD-Rag2(null) IL-2R?(null) (NR2G) mice similar to NOD-scidIL-2R?(null) (NOG) mice that are known as an excellent host to generate humanized mice. To evaluate the usefulness of NR2G mice as a host for humanized mice, the engraftment rates and differentiation of human cells after human hematopoietic stem cell (HSC) transplantation were compared among NR2G, NOG, and NOD-scid mice. For this purpose, the appropriate irradiation doses to expand the niche for human stem cells in the bone marrow were first determined. As a result, 8 and 2.5 Gy in adult, and 4 and 1 Gy in newborn NR2G and NOG mice, respectively, were found to be appropriate. Next, 5 × 10(4) human umbilical cord blood CD34(+) cells were intravenously inoculated into irradiated adult or newborn of the immunodeficient mice. These HSC transplantation experiments demonstrated that both NR2G and NOG mice showed high engraftment rates compared with NOD-scid mice, although NOG mice showed a slightly higher engraftment rate than that for NR2G mice. However, no difference was found in the human cell populations differentiated from HSCs between NR2G and NOG mice. The HSC transplantation experiments to adults and newborns of two immunodeficient mice also revealed that the HSC transplantation into newborn mice resulted in higher engraftment rate than those into adults. These results showed that NR2G mice could be used as an alternative host to NOG mice to generate humanized mice.
| S-EPMC4206736 | biostudies-literature
Project description:C3 glomerulopathy (C3GP) is a disease entity caused by abnormality of the complement alternative pathway (AP) and characterized by C3 deposition in glomeruli. Many variations or mutations of complement factors are believed to underlie the susceptibility to C3GP, but there is a lack of experimental evidence. We have recently reported a patient with C3 glomerulonephritis (C3GN) and compound heterozygosity of two novel variations in the complement factor (CFI). Here, we generated a mouse model to mimic the CFI variations for studying pathogenicity of CFI variations in C3GN development. We used the CRISPR/Cas9 system to make mutant mouse lines that carried D288G and P467S mutations in CFI, respectively, and crossed them to generate mice with compound heterozygosity of CFI D288G and P467S. The mice were all normal in either SPF (specific pathogen free) or regular environment. When treated with lipopolysaccharides (LPS), a bacterial endotoxin that mimics infection and sepsis, the mice developed albuminuria, kidney function impairment, and C3 glomerular deposition at levels comparable with the wild-type mice. The mice with other genotypes concerning CFI D288G and P467S were also tested in parallel. Unexpectedly, we found that the D288G homozygotes all developed severe mesangial deposition of C3 in the LPS model, indicating that CFI D288G variation was involved in the C3 deposition, a key feature of C3GN. The mouse lines generated in the present study can be used to further study the role of CFI variations in C3GN development; in addition, they may be used to screen and test infections and environmental factors capable of triggering C3GN.
| S-EPMC8209374 | biostudies-literature
Project description:Melanoma, an immunogenic tumor, is the first indication where oncolytic viruses are now becoming part of clinical practice. ONCOS-102, a transgened adenovirus, has shown to act as a primer of relevant tumor targeting immune cells both in preclinical and clinical melanoma studies. Strategies to augment its effectiveness warrant investigation. Combination therapy of ONCOS-102 with the checkpoint inhibitor (CPI) pembrolizumab was evaluated in a quasi-human animal model, the humanized NOG mouse model. A dosing schedule of the combination, beginning the CPI concurrently with the oncolytic viral therapy and continuing the CPI treatment, appeared to induce an abscopal effect in untreated tumor lesions. Concurrent combination therapy with checkpoint inhibitors may improve the induction of antitumor immune responses of ONCOS-102.
| S-EPMC6771875 | biostudies-other
Project description:BACKGROUND AND PURPOSE: Humanized mice for the nuclear receptor peroxisome proliferator-activated receptor ? (PPAR?), termed PPAR? knock-in (PPAR? KI) mice, were generated for the investigation of functional differences between mouse and human PPAR? and as tools for early drug efficacy assessment. EXPERIMENTAL APPROACH: Human PPAR? function in lipid metabolism was assessed at baseline, after fasting or when challenged with the GW0742 compound in mice fed a chow diet or high-fat diet (HFD). KEY RESULTS: Analysis of PPAR? mRNA levels revealed a hypomorph expression of human PPAR? in liver, macrophages, small intestine and heart, but not in soleus and quadriceps muscles, white adipose tissue and skin. PPAR? KI mice displayed a small decrease of high-density lipoprotein-cholesterol whereas other lipid parameters were unaltered. Plasma metabolic parameters were similar in wild-type and PPAR? KI mice when fed chow or HFD, and following physiological (fasting) and pharmacological (GW0742 compound) activation of PPAR?. Gene expression profiling in liver, soleus muscle and macrophages showed similar gene patterns regulated by mouse and human PPAR?. The anti-inflammatory potential of human PPAR? was also similar to mouse PPAR? in liver and isolated macrophages. CONCLUSIONS AND IMPLICATIONS: These data indicate that human PPAR? can compensate for mouse PPAR? in the regulation of lipid metabolism and inflammation. Overall, this novel PPAR? KI mouse model shows full responsiveness to pharmacological challenge and represents a useful tool for the preclinical assessment of PPAR? activators with species-specific activity.
| S-EPMC3171871 | biostudies-other
Project description:1. Humanized-liver mice, in which the liver has been repopulated with human hepatocytes, have been used to study aspects of human liver physiology such as drug metabolism, toxicology and hepatitis infection. However, the procurement of human hepatocytes is a major problem in producing humanized-liver mice because of the finite nature of the patient-derived resource. 2. In order to overcome this limitation, the human hepatic cell line HepaRG® were evaluated as promising donor cells for liver reconstitution in the TK-NOG mouse model. 3. We demonstrate that, in vivo, transplanted confluent culture or differentiated HepaRG® cells proliferated and differentiated toward both hepatocyte-like and biliary-like cells within the recipient liver. In contrast, proliferative HepaRG® cells could engraft TK-NOG mouse liver but could differentiate only toward biliary-like cells. The differentiation to hepatocyte-like cells was characterized by the detection of human albumin in the recipient mouse serum and was confirmed by immunohistochemical staining for human leukocyte antigen, human albumin, cytochrome P450 3A4, and multidrug resistance-associated protein 2. Biliary-like cells were characterized by positive staining for cytokeratin-19. 4. These results indicated that the differentiated HepaRG® cells are a possible cell source for generating humanized-liver mice, which are a useful model for in vivo studies of liver physiology.
| S-EPMC3906414 | biostudies-literature
Project description:BackgroundMost frequent red cell (RBC) donors and many first-time donors are iron deficient, but meet haemoglobin standards. However, the effects of donation-induced iron deficiency on RBC storage quality are unknown. Thus, we used a mouse model to determine if donor iron deficiency reduced post-transfusion RBC recovery.MethodsWeanling mice received a control diet or an iron-deficient diet. A third group receiving the iron-deficient diet was also phlebotomised weekly. This provided 3 groups of mice with different iron status: (1) iron replete, (2) mild iron deficiency with iron-deficient erythropoiesis, and (3) iron-deficiency anaemia. At ten weeks of age, blood was collected, leucoreduced, and stored at 4 ºC. After 12 days of storage, 24-hour (h) post-transfusion RBC recovery was quantified in recipients by flow cytometry.ResultsBefore blood collection, mean haemoglobin concentrations in the iron-replete, iron-deficient, and iron-deficiency anaemia donor mice were 16.5±0.4, 11.5±0.4, and 7.0±1.4 [g/dL± 1 standard deviation (SD)], respectively (p<0.01 for all comparisons between groups). The 24-h post-transfusion RBC recoveries in recipients receiving transfusions from these three cohorts were 77.1±13.2, 66.5±10.9, and 46.7±15.9 (% ±1 SD), respectively (p<0.05 for all comparisons between groups).DiscussionIn summary, donor iron deficiency significantly reduced 24-h post-transfusion RBC recovery in recipient mice. RBCs from mice with mild iron deficiency and iron-deficient erythropoiesis, with haemoglobin levels similar to those used for human autologous blood donation, had intermediate post-transfusion RBC recovery, as compared to iron-replete donors and those with iron-deficiency anaemia. This suggests that, in addition to the effects of iron deficiency on donor health, frequent blood donation, leading to iron-deficient erythropoiesis, may also have adverse effects for transfusion recipients.
| S-EPMC5336338 | biostudies-literature
Project description:Complement protein C3 is the pivotal component of the complement system. Previous studies have demonstrated that C3 has implications in various human diseases and exerts profound functions under certain conditions. However, the delineation of pathological and physiological roles of C3 has been hampered by the insufficiency of suitable animal models. In the present study, we applied the clustered regularly interspaced short palindromic repeat (CRISPR)/CRISPR-associated (Cas) system to target the C3 gene in porcine fetal fibroblasts. Our results indicated that CRISPR/Cas9 targeting efficiency was as high as 84.7%, and the biallelic mutation efficiency reached at 45.7%. The biallelic modified colonies were used as donor for somatic cell nuclear transfer (SCNT) technology to generate C3 targeted piglets. A total of 19 C3 knockout (KO) piglets were produced and their plasma C3 protein was undetectable by western blot analysis and ELISA. The hemolytic complement activity and complement-dependent cytotoxicity assay further confirmed that C3 was disrupted in these piglets. These C3 KO pigs could be utilized as a valuable large animal model for the elucidation of the roles of C3.
| S-EPMC5503937 | biostudies-literature