Name: huTNF(DBA/1) Mouse
Brand: サイヤジェン（Cyagen）
SKU: C001587

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huTNF(DBA/1) Mouse

製品名

huTNF(DBA/1) Mouse

製品ID

C001587

系統名

DBA/1-Tnf^em1(hTNF)/Cya

背景情報

DBA/1

状況

このマウス系統を論文で使用する場合は、「huTNF(DBA/1) Mouse（カタログ番号C001587）はサイアジェンから購入しました。」と引用してください。

HUGO-GT Humanized Models

Cytokine Gene Humanized Mouse Models

Rheumatoid Arthritis

製品タイプ

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遺伝子型

性別

数量

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HUGO-GT Humanized Models

Cytokine Gene Humanized Mouse Models

Rheumatoid Arthritis

基本情報

検証 Data

系統詳細

The tumor necrosis factor-alpha (TNF/TNF-α) gene encodes a pro-inflammatory cytokine belonging to the TNF superfamily. It is primarily produced by macrophages/monocytes during acute inflammation. TNF-α regulates immune cell function by binding to its receptors TNFRSF1A/TNFR1 and TNFRSF1B/TNFBR, participating in normal inflammatory and immune responses. TNF-α is involved in various biological processes, including cell proliferation, differentiation, apoptosis, lipid metabolism, and coagulation. This factor is associated with several diseases, such as autoimmune conditions, insulin resistance, psoriasis, rheumatoid arthritis, ankylosing spondylitis, tuberculosis, autosomal dominant polycystic kidney disease, and cancer. Mutations in the TNF-α gene impact susceptibility to cerebral malaria, septic shock, and Alzheimer’s disease^[1-2]. In mice, defects in this gene are associated with impaired responses to bacterial infections, defects in the organization of follicular dendritic cell networks and germinal centers, and a lack of primary B cell follicles.

The huTNF(DBA/1) mice is a mouse Tnf gene humanized model. The mouse Tnf gene in the DBA/1 strain is replaced with the human TNF gene, including the 5’UTR and 3’UTR. This model is useful for diseases researches such as rheumatoid arthritis (RA). Since immunization of DBA/1 mice with type II collagen leads to severe polyarthritis mediated by the autoimmune response, DBA/1 mice are widely used for RA model construction. Therefore, huTNF(DBA/1) mice constructed on a DBA/1 strain background can be used to research immune-related diseases such as RA. In addition, based on the independently developed TurboKnockout fusion BAC recombination technology, Cyagen can also generate mutation models based on this strain and provide customized services.

参考文献

Liu ZG. Molecular mechanism of TNF signaling and beyond. Cell Res. 2005 Jan;15(1):24-7.

Kalliolias GD, Ivashkiv LB. TNF biology, pathogenic mechanisms and emerging therapeutic strategies. Nat Rev Rheumatol. 2016 Jan;12(1):49-62.

Hou Y, Lin H, Zhu L, Liu Z, Hu F, Shi J, Yang T, Shi X, Zhu M, Godley BF, Wang Q, Li Z, Zhao Y. Lipopolysaccharide increases the incidence of collagen-induced arthritis in mice through induction of protease HTRA-1 expression. Arthritis Rheum. 2013 Nov;65(11):2835-46.

系統作製戦略

The mouse Tnf genome plus flanking sequence was replaced with the human TNF genome plus flanking sequence.

Figure 1. Schematic diagram of gene editing and targeting in huTNF(DBA/1) mice.

適用分野

Research on Rheumatoid Arthritis (RA);

Research on TNF-α signaling pathway;

Research on the pathogenesis and treatment of other TNF-α related diseases.

検証 Data

1. Expression of human TNF gene

The RT-qPCR results showed that there was significant expression of the human TNF gene in the thymus and spleen of huTNF(DBA/1) mice, while no expression of the human TNF gene was detected in DBA/1 wild-type mice. (ND: Not detected)

Figure 2. Detection of human TNF gene expression in the thymus and spleen of DBA/1 wild-type mice and huTNF(DBA/1) mice (10 weeks old, female ♀).

2. Expression of mouse Tnf gene

RT-qPCR results showed that the murine Tnf gene was expressed in both the thymus and spleen of DBA/1 mice, while there was no expression of the murine Tnf gene in huTNF(DBA/1) mice.

Figure 3. Detection of murine Tnf gene expression in the thymus and spleen of 10-week-old female DBA/1 wild-type mice and huTNF(DBA/1) mice.

3. Expression of TNF protein after LPS induction

The mice were intraperitoneally injected with LPS at a dose of 1.5 mg/kg to induce high expression of TNF protein**. Six hours later, the mouse serum was collected for ELISA detection. The results showed that after LPS induction, the human-derived TNF protein was significantly expressed in the serum of huTNF(DBA/1) mice, and the mouse-derived TNF protein was hardly expressed. After LPS induction in DBA/1 wild-type mice, the mouse-derived TNF protein was significantly expressed in the serum, while the content of human-derived TNF protein was extremely low.

Figure 4. ELISA detection of TNF protein expression in the serum of 10-week-old female DBA/1 wild-type mice and huTNF(DBA/1) mice (huTNF)*. *The human TNFα protein ELISA kit used for detection was from Thermo Fisher: Human TNF alpha ELISA Kit, Ultrasensitive (Catalog No.: KHC3014); The mouse TNFα protein ELISA kit used for detection was from MULTI SCIENCES: Mouse TNF-a ELISA Kit (EK282/4). **LPS is an endotoxin that can be used to stimulate the immune response in mice. LPS immunization can exacerbate the severity of arthritis ^[3].

4. Construction of Collagen-Induced Arthritis (CIA) Model with Type II Collagen for Pharmacodynamic Validation of Adalimumab.

All the mouse models used in the experiment are female.

Figure 5. Experimental Arrangement for CIA Model Construction and Adalimumab Treatment.

Figure 6. Images of Mouse Joints.

Figure 7. Growth Curves and Clinical Scores. The results show no significant difference in body weight between the model and control groups of huTNF(DBA/1) mice and DBA/1 mice. Clinical scores in the model groups of huTNF(DBA/1) and DBA/1 mice significantly increased, indicating successful CIA model construction. The disease progressed more rapidly and severely in the huTNF(DBA/1) model group compared to the DBA/1 model group. After Adalimumab treatment, the average clinical score of the huTNF-CIA+Adalimumab efficacy group was significantly lower than that of the model group, indicating effective disease control in mice (Note: Efficacy is related to treatment duration and dose).

Figure 8. Histopathology. ① No significant abnormalities were observed in the joint cavities of the control groups of huTNF(DBA/1) and DBA/1 mice, with no significant synovial hyperplasia and intact cartilage structure. (Blue: No significant synovial hyperplasia; Red: Normal joint cavity without deformation or fusion; Green: Intact cartilage surface without erosion or damage). ② The model group of DBA/1 mice showed significant synovial hyperplasia and thickening, with inflammatory cells invading the joint cavity. The synovium formed pannus and bone erosion, with excessive proliferation of synovial cells eroding cartilage and bone, resulting in the disappearance of the joint space. Pannus formation or extensive pannus was observed in multiple areas. ③ The model group of huTNF(DBA/1) mice showed synovial cell hyperplasia, proliferation of fibroblast-like cells and synovial phagocytes, accompanied by infiltration of inflammatory cells such as lymphocytes and plasma cells. The cartilage was affected by synovium, pannus, and inflammatory cells, with cartilage ulcer surfaces covered by connective tissue or fibrocartilage and invaded by new blood vessels. Bone tissue damage and joint cavity deformation were observed. ④ In the huTNF(DBA/1)-CIA+Adalimumab efficacy group, Adalimumab treatment significantly improved synovial hyperplasia and inflammatory infiltration; cartilage showed slight erosion (slightly uneven); joints were normal, with mild local cartilage defects and no pannus formation.

Figure 9. Pathological Score. The results show that pathological scores in the model groups (G2, G4) of huTNF(DBA/1) and DBA/1 mice were significantly higher than those in the control groups, with the disease severity being greater in the huTNF(DBA/1) model group. After Adalimumab treatment, synovial hyperplasia and inflammatory infiltration in arthritic mice were significantly improved, with slight cartilage erosion and mild local cartilage defects, resulting in significantly lower pathological scores.

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Collagen-induced arthritis (CIA) was established via subcutaneous injection of bovine type II collagen at the tail base to verify TNFi efficacy. CIA was successfully induced in both groups, and TNFi effectively alleviated arthritis in male huTNF(DBA/1) mice.

Figure 10. Arthritis scores of huTNF(DBA/1) mice in blank control and TNFi treatment groups (n=4, male). *These data were provided by Cyagen Biosciences partners.

6. Pharmacodynamic Validation of Infliximab

(A) Schematic diagram of the experimental procedure. On Day 0, a mixture of anti-type II collagen 5-clone monoclonal antibodies was injected to induce the CAIA model. On Day 3, LPS was administered to enhance inflammation. Infliximab was then injected intraperitoneally for treatment, and disease phenotypes were assessed throughout the experiment.

(B) Body weight changes during the experiment. Body weight trends were generally consistent between the treatment and control groups, indicating that Infliximab administration did not cause obvious systemic toxicity (mean ± SD).

(C–D) Hind paw swelling.

(E) Arthritis score. Following Infliximab treatment, both hind paw swelling and clinical arthritis scores were significantly lower in huTNF(DBA/1) mice compared with the control group, indicating effective suppression of inflammatory responses (mean ± SEM). Significance was determined by two-way ANOVA; unlabeled comparisons indicate no significant difference. huTNF(DBA/1)+CAIA vs. huTNF(DBA/1)+CAIA+Infliximab, **P < 0.01, ***P < 0.001.

The results demonstrate that Infliximab significantly alleviates CAIA-induced arthritis in huTNF(DBA/1) mice, validating the functional expression of human TNF. These findings confirm that inflammatory progression in the CAIA model is dependent on human TNF signaling, further verifying the successful functional expression of human TNF in huTNF(DBA/1) mice.

Figure 11. Infliximab efficacy validation protocol and results (8-week-old, female mice, n=5). *Infliximab is a chimeric anti-human TNF‑α monoclonal antibody that specifically neutralizes human TNF‑α with negligible activity against mouse TNF‑α, and is used for the treatment of autoimmune inflammatory diseases. *These data were provided by Cyagen Biosciences partners.

huTNF(DBA/1) Mouse

基本情報

検証 Data

関連リソース

遺伝子名

遺伝子別名

NCBI ID

染色体

MGI ID

さらに

データシート

系統詳細

参考文献

系統作製戦略

適用分野

1. Expression of human TNF gene

2. Expression of mouse Tnf gene

3. Expression of TNF protein after LPS induction

4. Construction of Collagen-Induced Arthritis (CIA) Model with Type II Collagen for Pharmacodynamic Validation of Adalimumab.

5. --

6. Pharmacodynamic Validation of Infliximab

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