KS (inducible) Mice

Catalog Number: C001514

Strain Name: C57BL/6JCya-Kras^{tm1(LSL-G12D)}Sftpc^{em2(IRES-MerCreMer)}/Cya

Genetic Background: C57BL/6JCya

Reproduction: Heterozygous LSL-K-ras G12D mice x Heterozygous/Homozygous Sftpc-MerCreMer mice

Note: Non-induced mice for this strain are the default shipment. Please get in touch with us if induced mice are required. The recommended induction protocol is included at the end of this manual.

Strain Description

KRAS is an oncogene that encodes the K-Ras protein, a key component of the RAS/MAPK signaling pathway. The K-Ras protein plays an important regulatory role in a variety of cellular processes, including cell growth, proliferation, maturation, and differentiation. KRAS gene mutations are a common cause of cancer, with approximately 30% of cancer patients harboring KRAS mutations. These mutations are predominantly single-nucleotide missense mutations, with over 80% occurring at the 12th amino acid residue (G12), particularly the KRAS G12D mutation, which is very common in lung and pancreatic cancers. The G12D mutation enhances the activity of the K-Ras protein, leading to uncontrolled cell growth and division, thereby promoting tumor formation. The K-Ras protein is highly conserved between mice and humans, with only differences at the 132nd and 187th amino acid residues ^[1–2].

The SFTPC gene encodes surfactant protein C (SP-C), one of the four major proteins that make up surfactant. Surfactant is a mixture of lipids and proteins that coats the surface of lung tissue, reducing respiratory resistance. It is produced and secreted by alveolar cells, and its main function is to maintain the stability of lung tissue by reducing the surface tension of lung fluid. In addition, SP-C protein is involved in lung development and function, including alveolar formation, airway remodeling, and immune defense. The SFTPC gene is expressed primarily in the lung, with the highest expression in the lower lobes, right lung, upper lobes, left upper lobes, and visceral pleura. It is also expressed at lower levels in other tissues. Type II alveolar cells are the main producers and secretors of surfactants, and the SFTPC gene is highly expressed in these cells, making it a specific marker of this cell type.

The KS (inducible) mouse is an induced lung cancer model that is constructed by crossing LSL-K-ras G12D mice (Catalog number: C001064), a conditional over-expressing K-Ras G12D mutant gene mouse strain, and Sftpc-MerCreMer mice (Catalog number: C001501), a type II alveolar cell-specific Cre recombinase expressing mouse strain, and then inducing with tamoxifen. Tamoxifen can trigger sequence recombination between loxP sites mediated by Cre recombinase in the type II alveolar cells of the offspring mice, resulting in the specific deletion of the Loxp-Stop-Loxp (LSL) gene silencing element in the type II alveolar cells, thereby enabling the K-Ras G12D mutant gene to be selectively expressed in the lung tissue.

Strain Strategy

Generated by crossing LSL-K-ras G12D mice with Sftpc-MerCreMer mice.

Application

KS (inducible) mice can be used to study the mechanisms of the occurrence and metastasis of non-small cell lung cancer (NSCLC), as well as the screening, development, and evaluation of therapeutic drugs.

Validation Data

1. Survival and growth curves

Figure 1. Survival and weight change of KS mice (n=10). KS mice at 4 weeks of age were induced to develop lung cancer by intraperitoneal injection of 75 mg/kg tamoxifen for 4 consecutive days. The survival rate and weight changes of the mice were monitored. (Note: All mice in subsequent experiments in this manual were induced with tamoxifen according to this protocol.) The results showed that the weight of KS mice significantly decreased after tamoxifen induction. The mice started to die after 9 weeks of induction. The mortality rate of KS mice reached 100% at 13 weeks after induction.

2. CT scan of the lungs

Figure 2. Lung CT scans of KS mice and wild-type mice at 6 weeks after tamoxifen induction. The results showed that KS mice treated with tamoxifen at 4 weeks of age developed obvious tumor tissue infiltration in the lungs 6 weeks after induction (the light-shaded area in the figure).

3. Morphology of the lungs (3 weeks after induction)

Figure 3. Lung morphology of female and male KS mice 3 weeks after tamoxifen induction. Tamoxifen induction was carried out in KS mice at 4 weeks of age, and the mice were dissected 3 weeks after completion of induction. The results showed that the lungs of KS mice in the tamoxifen-induced group showed significant hyperplasia and enlargement at 3 weeks after completion of induction compared with those of KS mice in the non-induced group.

4. Morphology of the lungs and spleen (6, 9 and 12 weeks after induction)

Figure 4. Morphology of the lungs and spleens of female KS mice and wild-type control mice at 6, 9, and 12 weeks after tamoxifen induction. The results showed that KS mice treated with tamoxifen at 4 weeks of age developed tumor tissue growth and invasion in the lungs 6 weeks after induction, with the lungs showing a dense and enlarged structure. At 12 weeks after induction, the spleens of KS mice showed abnormal hyperplasia.

5. Hematoxylin and Eosin (H&E) staining

1) Lung H&E staining

Figure 5. The H&E staining results of lung tissue from female KS mice treated with tamoxifen or corn oil at 6 and 12 weeks. The results showed that, compared with the control group, the tamoxifen-treated KS mice had obvious infiltration in the lungs. After 6 weeks of tamoxifen induction, the lungs of KS mice had already begun to show obvious cancerization. The lesions were further aggravated after 12 weeks of induction.

2) H&E staining of other tissues

Figure 6. The H&E staining results of kidneys, liver, spleen, and pancreas from female KS mice treated with tamoxifen or corn oil at 12 weeks. The results showed that compared with the control group, the kidneys, liver, and pancreas of KS mice treated with tamoxifen for 12 weeks were normal, while the spleen showed abnormal hyperplasia of white pulp.

Recommended Induction Protocol

Injection Solution Preparation: Thoroughly mix ethanol and corn oil in a 1:9 ratio to prepare the injection solution.

Tamoxifen Dissolution: Dissolve tamoxifen in the injection solution to a final concentration of 10mg/mL. If tamoxifen is insoluble or difficult to dissolve, sonication can be used to assist dissolution and the sonication time can be appropriately increased. After the preparation is complete, the solution should be stored in a light-protected manner.

Tamoxifen Dosage: Administer tamoxifen induction at a dose of 75mg/kg/day for 4 consecutive days, based on the actual weight of the mice.

Administration Method: Intraperitoneal injection (i.p).

Starting Age: 4 weeks to 8 weeks of age.

Reagent Source: Tamoxifen (T5648-1G, Sigma, Burlington, MA, US); Corn Oil (C116025, Shanghai Aladdin Biochemical Technology, Shanghai, CN); Anhydrous Ethanol (10009218, Sinopharm Group, Shanghai, CN).

Table 1. Recommended tamoxifen induction doses and regimens*

*Recommended based on internal experimental results, the final induction effect may vary with the actual living environment of the mice and the reagents used, and is provided here for reference only.

References

[1] Li S, Balmain A, Counter CM. A model for RAS mutation patterns in cancers: finding the sweet spot. Nat Rev Cancer. 2018 Dec;18(12):767-777.

[2] Filiberti A, Ventafridda V, Costa A. What is the best treatment for early breast cancer? A psychosocial answer. Ann Oncol. 1995 May;6(5):417-9.