Gene target for predicting and / or diagnosing and / or improving osteoporosis and application thereof

Abstract

The invention relates to a gene target for predicting and / or diagnosing and / or improving osteoporosis and application thereof, and belongs to the technical field of biological medicine. The invention provides a gene target for predicting and / or diagnosing and / or improving osteoporosis, the gene target is CYP7B1, and the amino acid sequence of the CYP7B1 is as shown in SEQ ID NO. 1. Cell experiments and animal experiments prove that the expression level change of CYP7B1 is closely related to osteoporosis, and CYP7B1 can be used as a target for prediction or auxiliary diagnosis of osteoporosis. Overexpression of the gene target shows that osteoporosis related indexes are remarkably improved, and the compound or the derivative thereof capable of correspondingly improving the expression level of the gene target can be selected to be applied to preparation of medicines for improving osteoporosis.

Description

Technical Field

[0001] This invention relates to the field of biomedical technology, and in particular to a gene target for predicting and / or diagnosing and / or improving osteoporosis and its application. Background Technology

[0002] Osteoporosis (OP) is a systemic skeletal disease characterized by decreased bone mass and deterioration of bone microstructure, leading to increased bone fragility and susceptibility to fractures. Osteoporosis has become an undeniable global public health problem. According to the International Osteoporosis Foundation, in 2010, the number of people aged 50 and older at high risk of osteoporotic fractures worldwide was 158 million, and this number is projected to double by 2040. The functional impacts of osteoporosis-related fractures include pain, dependency, depression, skeletal deformities, and impairment of basic daily activities. Fractures associated with osteoporosis of the hip, vertebrae, and pelvis are common causes of morbidity and mortality in the elderly. Many risk factors contribute to osteoporosis, such as aging, reduced mechanical stimulation, hormonal imbalances, and unhealthy lifestyle habits. These factors can disrupt the dynamic balance between osteoblast-mediated bone formation and osteoclast-driven bone resorption, the imbalance between osteogenic and adipogenic differentiation of mesenchymal stem cells, and oxidative stress-induced cellular DNA damage, apoptosis, and aging.

[0003] Bone marrow mesenchymal stem cells (BM-MSCs), originating from the mesoderm, are adult stem cells capable of self-renewal and multi-lineage differentiation into adipocytes, osteoblasts, and chondrocytes, playing a crucial role in maintaining bone homeostasis. Increasing evidence suggests that alterations in the number and function of mesenchymal stem cells are a significant contributing factor to osteoporosis. However, the underlying molecular mechanisms remain unclear.

[0004] Current drug treatments for osteoporosis primarily consist of anti-resorption drugs and anabolic agents. Anti-resorption drugs mainly work by inhibiting osteoclast recruitment and activity, thereby reducing the rate of bone remodeling and reversing temporary defects caused by resorption cavities that have not yet formed or completed, allowing for moderate bone resorption; examples include bisphosphonates and denosumab. Anabolic agents, on the other hand, mainly achieve their effect by altering bone remodeling, bone shaping, or a combination of both, such as teriparatide. Compared to anti-resorption drugs, anabolic agents have better anti-fracture efficacy and can increase bone mineral density to a greater extent. The effects of anabolic agents are transient, thus requiring a transition to anti-resorption drugs. The optimal strategy for cycling anabolic agents, anti-resorption drugs, and the withdrawal period remains to be determined. Summary of the Invention

[0005] The purpose of this invention is to overcome the shortcomings of the prior art and provide a new gene target for predicting and / or diagnosing and / or treating osteoporosis and its application in the preparation of products for predicting and / or diagnosing and / or improving osteoporosis.

[0006] To achieve the above objectives, the technical solution adopted by the present invention is as follows: In a first aspect, the present invention provides a gene target for predicting and / or diagnosing and / or improving osteoporosis, the gene target being CYP7B1, the amino acid sequence of which is shown in SEQ ID NO.1.

[0007] This invention demonstrates through cell and animal experiments that changes in CYP7B1 expression levels are closely related to osteoporosis, and that CYP7B1 can serve as a target for predicting or assisting in the diagnosis of osteoporosis.

[0008] Secondly, the present invention provides the application of the above-mentioned gene targets in the preparation of products for predicting osteoporosis.

[0009] Thirdly, the present invention provides the application of the above-mentioned gene targets in the preparation of products for diagnosing osteoporosis.

[0010] As a preferred embodiment of the application described in this invention, the product includes reagents involved in the technology for detecting the expression level of CYP7B1 in isolated biological samples.

[0011] In a preferred embodiment of the application described in this invention, the ex vivo biological sample is bone marrow mesenchymal stem cells.

[0012] As a preferred embodiment of the application described in this invention, the techniques for detecting the expression level of CYP7B1 in ex vivo biological samples include, but are not limited to, at least one of: RT-PCR, real-time quantitative PCR, immunological detection, in situ hybridization, microarray, high-throughput sequencing, chromatography, mass spectrometry, and gel electrophoresis.

[0013] As a preferred embodiment of the application described in this invention, the RT-PCR and real-time quantitative PCR include PCR primers, the nucleotide sequences of which are shown in SEQ ID NO.2-5.

[0014] As a preferred embodiment of the application described in this invention, the product includes at least one of food, health products, pharmaceuticals, and testing supplies.

[0015] Fourthly, a kit for predicting and / or diagnosing osteoporosis, comprising reagents for detecting the aforementioned gene targets.

[0016] As a preferred embodiment of the kit described in this invention, the reagent for detecting the above-mentioned gene targets is mainly at least one of the following: RT-PCR, real-time quantitative PCR, immunological detection, in situ hybridization, microarray, high-throughput sequencing, chromatography, mass spectrometry, and gel electrophoresis.

[0017] As a preferred embodiment of the kit described in this invention, the RT-PCR and real-time quantitative PCR include PCR primers, the nucleotide sequences of which are shown in SEQ ID NO.2-5.

[0018] Fifthly, the present invention provides the application of the above-mentioned gene targets as detection targets in screening drugs to improve osteoporosis.

[0019] Sixthly, the present invention provides the application of compounds that enhance the expression level of the above-mentioned gene targets in the preparation of products that improve osteoporosis.

[0020] This invention, through CYP7B1 overexpression modification of bone marrow mesenchymal stem cells, found that it can inhibit the adipogenic differentiation ability of bone marrow mesenchymal stem cells derived from osteoporosis patients, thereby achieving the effect of improving osteoporosis.

[0021] As a preferred embodiment of the application described in this invention, the product includes at least one of food, pharmaceuticals, and health products.

[0022] As a preferred embodiment of the application described in this invention, the compound that enhances the expression level of the above-mentioned gene target includes a lentivirus that overexpresses CYP7B1.

[0023] In a seventh aspect, the present invention provides a medicine for improving osteoporosis, comprising a compound or a derivative thereof that increases the expression level of the aforementioned gene target.

[0024] In a preferred embodiment of the pharmaceutical product of the present invention, the pharmaceutical product further includes pharmaceutically acceptable excipients.

[0025] Eighthly, the present invention provides a method for constructing a mouse model of osteoporosis, comprising the following steps: by... Cyp7b1 fl / fl mice and Prx1-Cre Mice were hybridized to obtain Prx1-Cre ; Cyp7b1 fl / + Heterozygous mice, males Prx1-Cre ; Cyp7b1 fl / + Heterozygous mice and Cyp7b1 fl / fl Mouse backcrossing yielded the target genotype as Prx1-Cre ; Cyp7b1 fl / fl The mice, the Prx1-Cre ; Cyp7b1 fl / fl The mice in question are the osteoporosis mouse model.

[0026] Compared with the prior art, the beneficial effects of the present invention are as follows: (1) This invention has demonstrated through cell experiments and animal experiments that changes in the expression level of CYP7B1 are closely related to osteoporosis, and that CYP7B1 can be used as a target for predicting or assisting in the diagnosis of osteoporosis.

[0027] (2) The present invention found that the osteoporosis-related indicators were significantly improved by overexpressing the gene target. Compounds or their derivatives that can improve the expression level of the gene target can be selected and used in the preparation of materials to improve osteoporosis. Attached Figure Description

[0028] Figure 1 The results of flow cytometry analysis of human bone marrow mesenchymal stem cells in this invention are shown. Figure 2 The results of flow cytometry analysis of the mouse-derived bone marrow mesenchymal stem cells of this invention are shown below. Figure 3 The results of CYP7B1 expression level detection in mouse bone marrow mesenchymal stem cells in Example 1 of this invention are shown in Figure A, where A is the RNA expression level detection result, B is the protein expression level detection result, Sham is the sham-operated group, OVX is the postmenopausal osteoporosis model group, Young is the young mouse group, and Old is the elderly osteoporosis model group. Figure 4 The results of CYP7B1 expression level detection in human bone marrow mesenchymal stem cells in Example 1 of the present invention are shown. A represents the RNA expression level detection result, B represents the protein expression level detection result, Non-OP represents healthy individuals, and OP represents osteoporosis patients. Figure 5 The results of adipogenic differentiation detection of human bone marrow mesenchymal stem cells in Example 2 of the present invention are shown. In this example, A represents cells with CYP7B1 knockdown, B represents cells with CYP7B1 overexpression, Si-Ctrl represents the knockdown control group, Si-CYP7B1 represents the CYP7B1 knockdown group, Oe-Ctrl represents the overexpression control group, Oe-CYP7B1 represents the CYP7B1 overexpression group, Non-OP represents healthy individuals, and OP represents patients with osteoporosis. Figure 6 The images show the femoral bone mass detection results of mice under different treatments in Example 3 of this invention, where A is a CT scan image and B is related indices of femoral trabeculae and cortical bone. Cyp7b1 fl / fl The control group consisted of mice. Cyp7b1 fl / fl Prx1-cre To knock out CYP7B1 mice In the above figure, there is a mark marked " "" indicates a significant difference between the two groups (P < 0.05), marked with " "" indicates a significant difference between the two groups (P < 0.01), marked with " “” indicates a significant difference between the two groups (P < 0.001), while “ns” indicates no significant difference between the two groups (P > 0.05). Detailed Implementation

[0029] To better illustrate the purpose, technical solution, and advantages of the present invention, the present invention will be further described below in conjunction with specific embodiments.

[0030] Unless otherwise specified, all other materials and reagents used in the examples, comparative examples, and effect examples are commercially available.

[0031] The human bone marrow mesenchymal stem cells in the following examples were obtained from clinical departments and collected from osteoporosis patients and healthy individuals. The healthy individuals were obtained from outpatient health check-up participants. Both osteoporosis patients and healthy individuals signed informed consent forms.

[0032] The method for isolating human bone marrow mesenchymal stem cells (BMSCs) was as follows: Samples were obtained from bone marrow aspiration fluid from the donor's posterior superior iliac crest under aseptic conditions and cultured in DMEM incomplete medium supplemented with 10% (v / v) fetal bovine serum at 37°C. The culture medium was changed every 2-3 days. When the cell confluence reached 80%-90%, the cells were passaged at a 1:2 ratio using trypsin digestion or seeded into 12-well plates for subsequent experiments. Flow cytometry was used to detect the purity of the BMSCs. The results are as follows: Figure 1 As shown, CD29, CD73, and CD105 were positive in human bone marrow mesenchymal stem cells, while CD45, CD34, and HLA-DR were negative, indicating that human bone marrow mesenchymal stem cells were successfully isolated.

[0033] The mouse bone marrow mesenchymal stem cells used in the following examples were derived from postmenopausal osteoporosis models and aged osteoporosis models, while the control group bone marrow mesenchymal stem cells were derived from the sham-operated group and young mice. All animal experimental procedures were performed in accordance with the "Guidelines for the Care and Use of Laboratory Animals (8th Edition)" and were approved by the Laboratory Animal Management and Use Committee.

[0034] The above-mentioned postmenopausal osteoporosis model was established as follows: 8-week-old female C57BL / 6J mice underwent bilateral ovariectomy, while the control group received sham surgery. The mice were then fed for 8 weeks post-surgery before subsequent experiments.

[0035] The above-mentioned method for constructing the osteoporosis model in the elderly is as follows: mice are raised to 18 months of age, while young mice are raised to 8 weeks of age before subsequent experiments.

[0036] The method for isolating mouse bone marrow mesenchymal stem cells (BMSCs) was as follows: Under aseptic conditions, the femur and tibia of mice were dissected, surrounding soft tissues were removed, and bone marrow was flushed from the bone marrow cavity using a syringe in α-MEM complete medium (containing 10% (v / v) fetal bovine serum and 100 IU / ml penicillin-streptomycin). The collected cell suspension was seeded into culture flasks and cultured at 37°C and 5% CO2, with the medium changed every 3 days. When the cell confluence reached 80%-90%, the mouse BMSCs were passaged into two new culture flasks or seeded into 12-well plates for subsequent experiments using trypsin digestion. The purity of the BMSCs was determined by flow cytometry. The results are shown below. Figure 2 As shown, mouse-derived bone marrow mesenchymal stem cells were positive for CD44, CD105, and Sca1, while negative for CD45, CD11b, and CD34, indicating that mouse-derived bone marrow mesenchymal stem cells were successfully isolated.

[0037] The formulation of the adipogenic differentiation induction medium was: 10% FBS DMEM, 1 μM dexamethasone, 10 μg / mL insulin, 0.5 mM isobutyl methyl xanthine (IBMX), 0.2 mM indomethacin and 100 IU / mL penicillin-streptomycin.

[0038] The amino acid sequence of CYP7B1 is MAGEVSAATGRFSLERLGLPGLALAAALLLLALCLLVRRTRRPGEPPLIKGWLPYLGVVLNLRKDPLRFMKTLQKQHGDTFTVLLGGKYITFILDPFQYQLVIKNHKQLSFRVFSNK LLEKAFSISQLQKNHDMNDELHLCYQFLQGKSLDILLESMMQNLKQVFEPQLLKTTSWDTAELYPFCSSIIFEITFTTIYGKVIVCDNNKFISELRDDFLKFDDKFAYLVSNIPIELLGNVKSIREKIIKC FSSEKLAKMQGWSEVFQSRQDVLEKYYVHEDLEIGAHHLGFLWASVANTIPTMFWAMYYLLRHPEAMAAVRDEIDRLLQSTGQKKGSGFPIHLTREQLDSLICLESSIFEALRLSSYSTTIRFVEEDLTLS SETGDYCVRKGDLVAIFPPVLHGDPEIFEAPEEFRYDRFIEDGKKKTTFFKRGKKLKCYLMPFGTGTSKCPGRFFALMEIKQLLVILLTYFDLEIIDDKPIGLNYSRLLFGIQYPDSDVLFRYKVKS (SEQ ID NO.1).

[0039] Example 1 To verify the effect of CYP7B1 gene target expression level on osteoporosis, the expression level of CYP7B1 in extracted human and mouse bone marrow mesenchymal stem cells was detected by qRT-PCR and Western blotting.

[0040] RNA was extracted and reverse transcribed according to the instructions of the RNA extraction kit and reverse transcription kit. The qRT-PCR reaction was then performed according to the kit's formulation. The qRT-PCR reaction conditions were: 95℃ for 30s, one cycle; 95℃ for 5s, 60℃ for 30s, 40 cycles. The primers used are as follows: Human-derived qPCR-F 5'-AAAACCACAAGTTGGGACACG-3' (SEQ ID NO.2), qPCR-R 5'-GAAGCTCAATGGGTATGTTGGAT-3' (SEQ ID NO.3); Mouse-derived qPCR-F5'-GCTTGGTCTGCCTGGAAAGC-3' (SEQ ID NO.4) and qPCR-R5'-TATCCTCCTGCACTTCTCGGA-3' (SEQ ID NO.5).

[0041] According to 2 -ΔΔCT The relative expression level of CYP7B1 was calculated using the method described in the following figure. Figure 3-4 .

[0042] Western blot analysis was performed using the following method: 1) Take protein lysis buffer and treat bone marrow mesenchymal stem cells separately. Determine the protein concentration of each protein sample by BCA protein concentration assay. Add 5× SDS-PAGE protein loading buffer to the collected protein samples, heat at 100℃ for 10 min, and the protein loading amount is 20 μg. 2) 10% SDS-PAGE electrophoresis: The stacking gel was electrophoresed at a constant voltage of 90 V for 30 min, and the separating gel was electrophoresed at a constant voltage of 120 V for 100 min when the bromophenol blue entered the separating gel; 3) Transferring PVDF membranes: The transfer tank was placed in ice water, the transfer current was 300 mA, and the transfer time was 60 min. 4) Immediately seal with 5% skim milk for 60 minutes; wash the membrane with TBST 3 times, 5 minutes each time; 5) According to the primary antibody's instructions, dilute the primary antibody with 5% skim milk and incubate overnight at 4°C. The next day, wash the membrane three times with TBST for 5 minutes each time. According to the secondary antibody's instructions, dilute the secondary antibody with TBST and incubate at low speed on a shaker at room temperature for 1 hour. Wash the membrane three times with TBST for 5 minutes each time, and detect protein expression levels. The results are shown in […]. Figure 3-4 .

[0043] like Figure 3-4 As shown, the RNA and protein expression levels of CYP7B1 isolated from human and mouse bone marrow mesenchymal stem cells with osteoporosis showed a down-regulated trend. However, the RNA and protein expression levels of CYP7B1 in bone marrow mesenchymal stem cells isolated from healthy individuals, young mice, and sham-operated mice were significantly higher than those in bone marrow mesenchymal stem cells from patients with osteoporosis and mice. This indicates that there is a certain correlation between the down-regulation of CYP7B1 expression in bone marrow mesenchymal stem cells and osteoporosis.

[0044] Example 2 Enhanced adipogenic differentiation capacity of bone marrow mesenchymal stem cells is one of the important contributing factors to osteoporosis. To evaluate the effect of CYP7B1 on adipogenic differentiation of bone marrow mesenchymal stem cells, CYP7B1 was overexpressed or knocked down, as follows: Human bone marrow mesenchymal stem cells from healthy individuals and osteoporosis patients were seeded into 12-well cell culture plates at a density of 6 × 10⁶ cells / well. 4 Cells / well, cell confluence reached approximately 50%-70% the day after seeding. Bone marrow mesenchymal stem cells were transfected with CYP7B1 overexpression and knockdown siRNA.

[0045] To obtain CYP7B1 overexpressing lentivirus (Oe-CYP7B1), the pSLenti-EF1-EGFP-P2A-Puro-CMV-CYP7B1-3×FLAG-WPRE vector was used as the packaging plasmid. The human CYP7B1 coding sequence was obtained by PCR amplification, and a 3×FLAG tag was introduced at the C-terminus. The amplified fragment was ligated to the linearized pSLenti-EF1-EGFP-P2A-Puro-CMV backbone using NEBuilder HiFi, and then transformed into DH5α competent cells. Positive clones were selected for plasmid extraction, and the correctness of the inserted sequence was confirmed by Sanger sequencing. When constructing the empty vector control virus (Oe-Ctrl), the same pSLenti-EF1-EGFP-P2A-Puro-CMV vector was used, but without the insertion of the CYP7B1 fragment. Subsequently, packaging plasmids, packaging system helper plasmids (psPAX2, pMD2.G), and Oe-CYP7B1 or Oe-Ctrl plasmids were co-transfected into 293T cells to obtain lentiviral particles. Before transfection, the original culture medium was replaced with fresh antibiotic-free medium. Then, the prepared Oe-CYP7B1 or Oe-Ctrl lentiviral stock solution was diluted appropriately with serum-free medium to the optimal multiple of infection (MOI=40). The culture medium was replaced 24 hours after lentiviral transfection, and the induction medium was replaced 6 hours after siRNA transfection. The CYP7B1 siRNA sequence is 5′-GGUGGAAAGUACAUAACAUTT-3′.

[0046] After knockdown or overexpression of bone marrow mesenchymal stem cells, adipogenic differentiation induction medium was added for induction, with the medium changed every 3 days. Adipogenic levels were assessed on day 10 of induction. Oil Red O staining and quantitative analysis were used to detect adipogenic differentiation. Results are shown below. Figure 5 .

[0047] like Figure 5As shown, Oil Red O staining and quantitative analysis revealed the adipogenic differentiation of CYP7B1 knockdown and overexpression in bone marrow mesenchymal stem cells (BMSCs) from healthy individuals and osteoporosis patients. After CYP7B1 knockdown, the adipogenic differentiation capacity of BMSCs in both healthy individuals and osteoporosis patients was enhanced, and the degree of adipogenic differentiation was almost identical in both groups. However, after CYP7B1 overexpression, the adipogenic differentiation capacity of BMSCs in both osteoporosis patients and healthy individuals decreased, and the degree of adipogenic differentiation was similar between osteoporosis patients and healthy individuals, with no significant difference. This indicates that the expression level of CYP7B1 is closely related to the adipogenic differentiation capacity of BMSCs, and this correlation is negative. Increasing the expression level of CYP7B1 can effectively prevent adipogenic differentiation of BMSCs, thereby improving osteoporosis.

[0048] Example 3 To assess the association between abnormal CYP7B1 expression levels and osteoporosis in animals, CYP7B1 knockout mice were constructed for observation. The specific protocol is as follows: First, build Cyp7b1 fl / fl Mice (constructed by Shanghai Southern Model Biotechnology Co., Ltd.) and Prx1- Cre Mice (purchased from Jackson Laboratory). Subsequently, Cyp7b1 fl / fl mice and Prx1-Cre Mouse hybridization, resulting in Prx1- Cre ; Cyp7b1 fl / + Heterozygous mice. Males... Prx1-Cre ; Cyp7b1 fl / + mice and Cyp7b1 fl / fl Mouse backcrossing ultimately yielded the target genotype. Prx1-Cre ; Cyp7b1 fl / fl The mice were used for genotyping. Genomic DNA was extracted from CKO mice and PCR analysis was performed. The primer sequences used for genotyping are shown in Table 1.

[0049] Table 1. Primer sequences for genotyping Mouse femurs were harvested and subjected to Micro-CT scans. Related parameters of femoral trabeculae and cortical bone were analyzed, including bone volume fraction, trabecular thickness, cortical thickness, trabecular pattern factor, trabecular number, trabecular separation, and bone surface area / bone volume ratio. Results Figure 6 And Table 2.

[0050] Table 2. Related indices of femoral trabeculae and cortical bone (x̅±SD) like Figure 6 As shown, compared with the control group mice ( Cyp7b1 fl / fl Compared to mice, CYP7B1-specific knockout mice showed significantly reduced bone volume fraction, trabecular thickness, cortical thickness, trabecular pattern factor, and trabecular number, while significantly increased trabecular separation and bone surface area / bone volume. The significantly reduced bone mass in CYP7B1-specific knockout mice indicates a successful establishment of an osteoporosis mouse model, and that CYP7B1 deficiency is a crucial factor in the development of osteoporosis.

[0051] Finally, it should be noted that the above embodiments are only used to illustrate the technical solutions of the present invention and are not intended to limit the scope of protection of the present invention. Although the present invention has been described in detail with reference to preferred embodiments, those skilled in the art should understand that modifications or equivalent substitutions can be made to the technical solutions of the present invention without departing from the essence and scope of the technical solutions of the present invention.

Claims

1. A gene target for predicting and / or diagnosing and / or improving osteoporosis, characterized in that, The gene target is CYP7B1, and the amino acid sequence of CYP7B1 is shown in SEQ ID NO.

1.

2. The application of the gene target as described in claim 1 in the preparation of products for predicting osteoporosis.

3. The application of the gene target as described in claim 1 in the preparation of products for diagnosing osteoporosis.

4. The application as described in any one of claims 2-3, characterized in that, The product includes reagents used in techniques for detecting CYP7B1 expression levels in ex vivo biological samples.

5. The application as described in any one of claims 2-3, characterized in that, The techniques for detecting CYP7B1 expression levels in ex vivo biological samples include, but are not limited to, at least one of the following: RT-PCR, real-time quantitative PCR, immunological detection, in situ hybridization, microarray, high-throughput sequencing, chromatography, mass spectrometry, and gel electrophoresis.

6. A kit for predicting and / or diagnosing osteoporosis, characterized in that, Includes reagents for detecting the gene target described in claim 1.

7. The application of the gene target as described in claim 1 as a detection target in screening drugs to improve osteoporosis.

8. The use of the compound that improves the expression level of the gene target as described in claim 1 in the preparation of products that improve osteoporosis.

9. A medicine for improving osteoporosis, characterized in that, This includes compounds or derivatives thereof that increase the expression level of the gene target described in claim 1.

10. A method for constructing a mouse model of osteoporosis, characterized in that, Includes the following steps: Will Cyp7b1 fl / fl mice and Prx1-Cre Mice were hybridized to obtain Prx1-Cre ; Cyp7b1 fl / + Heterozygous mice, males Prx1-Cre ; Cyp7b1 fl / + Heterozygous mice and Cyp7b1 fl / fl Mouse backcrossing yielded the target genotype as Prx1-Cre ; Cyp7b1 fl / fl The mice, the Prx1-Cre ; Cyp7b1 fl / fl The mice in question are the osteoporosis mouse model.

Images