Application of bone-strengthening and blood-nourishing oral liquid in preparation of medicine for treating osteoporotic fracture

Abstract

The invention relates to the technical field of biological treatment of bone metabolic diseases, and discloses an application of a bone-strengthening and blood-generating oral liquid in preparation of a medicine for treating osteoporotic fracture.The application includes the steps that when the medicine is prepared, active components in the bone-strengthening and blood-generating oral liquid are combined with a Wnt signal transduction complex in osteoblasts in a targeted mode, and the bone-strengthening and blood-generating oral liquid is obtained; the phosphorylation activity of glycogen synthase kinase-3 is inhibited, so that-catenin enters a nucleus, Runx2 expression is started, and the synthesis rate of the N-terminal propeptide of the type I procollagen is increased; according to the method, a positive anabolism window is established in an original callus formation period of fracture healing by constructing an osteogenesis and osteoclast activity two-way decoupling biological regulation mechanism, so that a positive anabolism effect is achieved, and a positive anabolism effect is achieved. Bone metabolism pathological deviation in an osteoporosis state is corrected, secondary absorption of callus is inhibited while high osteogenic activity is maintained, and biomechanical fracture load of bones after fracture healing is improved.

Description

Technical Field

[0001] This invention relates to the application of a bone-strengthening and blood-nourishing oral liquid in the preparation of drugs for treating osteoporotic fractures, belonging to the field of biotherapy technology for bone metabolic diseases. Background Technology

[0002] Osteoporotic fractures are a serious complication of osteoporosis, commonly seen in the elderly. The healing process involves a complex dynamic balance between osteogenic and osteoclast activities. Current treatment strategies are mainly divided into two categories: anti-bone resorption and bone formation promotion. Bisphosphonates reduce bone loss by inhibiting osteoclast activity and are used for long-term osteoporosis management. Parathyroid hormones promote new bone formation by activating osteoblasts and are used to treat severe osteoporosis. Clinical practice usually involves internal fixation surgery combined with anti-osteoporosis drug intervention to maintain overall bone mass while promoting local fracture healing. In response to the specific traumatic microenvironment of osteoporotic fractures, existing drug therapies face the challenge of reconciling the pathological metabolic contradictions. While bone resorption inhibitors effectively inhibit osteoclasts, they also disrupt normal intercellular coupling signals, thereby inhibiting osteogenic activity and leading to an excessively reduced bone turnover rate. The frozen bone effect delays the ossification of cartilage and callus remodeling at the fracture site. Although osteogenic drugs can temporarily increase the rate of bone formation, long-term or improper use can easily induce a synchronous upregulation of osteoclast activity, resulting in secondary resorption in the callus area, increasing the risk of insufficient mechanical strength of bone structure after healing and refracture.

[0003] Although some studies have extracted specific plant monomers to intervene in this process, single-target regulation strategies are still insufficient in the face of complex bone metabolic networks. For example, Chinese invention patent CN111494356A discloses the application of salvianolic acid B in the preparation of drugs for treating osteoporosis and osteoporotic fractures. This technical solution confirms that salvianolic acid B can upregulate the expression of Runx2 protein and osteocalcin gene, and promote the osteogenic differentiation potential of bone marrow mesenchymal stem cells. However, the solution essentially relies on the unidirectional activation of a single osteogenic pathway and lacks a systemic mechanism for synchronous and specific inhibition of osteoclast activity. Under the pathological conditions of osteoporotic fractures, if only the osteogenic rate is increased, the imbalance of the RANKL / OPG signal axis in the microenvironment cannot be corrected. The newly formed callus is easily degraded rapidly due to the excessive activity of osteoclasts, resulting in the inability to build a stable positive anabolic window at the fracture site. This makes it difficult to fundamentally solve the clinical problems of poor callus quality and delayed recovery of biomechanical strength.

[0004] Therefore, breaking the physiological coupling dependence of bone metabolism and constructing a positive anabolic metabolic window of high osteogenicity and low resorption during the critical period of fracture healing is the technical problem to be solved by this invention. Summary of the Invention

[0005] To address the problems mentioned in the background art, the technical solution of the present invention is as follows: The application of a bone-strengthening and blood-nourishing oral liquid in the preparation of a drug for treating osteoporotic fractures. The bone-strengthening and blood-nourishing oral liquid is made from bone fluid, astragalus, codonopsis, ganoderma, jujube, and black fungus. When used in the preparation of the drug, the drug is limited to regulating the bone metabolic microenvironment at the fracture site through a bidirectional decoupling mechanism. This bidirectional decoupling mechanism is configured to quantitatively improve the quality of fracture healing through the following molecular biological pathways: The active ingredient in the drug targets and binds to the Wnt signaling complex within osteoblasts, inhibiting glycogen synthase kinase-3. Phosphorylation activity, blocking The degradation of α-catenin protein promotes its entry into the nucleus, which in turn binds to T-cell kinases and lymphocyte-enhancing factors to initiate the expression of Runx2 transcription factor, thereby increasing the synthesis rate of type I procollagen N-terminal propeptide in serum, a key indicator of bone formation, and driving the transformation of cartilaginous callus into bony callus. Simultaneously, the bidirectional uncoupling mechanism utilizes the paracrine effect generated by osteoblasts in response to activation of the Wnt signaling transduction complex to upregulate the synthesis and secretion of osteoprotegerin; osteoprotegerin competitively binds to nuclear factors in the extracellular microenvironment. B receptor activator ligand, blocking the binding of the ligand to the RANK receptor on the surface of osteoclast precursors, inhibits intracellular NF-κB in osteoclasts. Activation of the B signaling pathway reduces the expression level of tartrate-resistant acid phosphatase, a key indicator of bone resorption, in serum. Based on the increased concentration of type I procollagen N-terminal propeptide and the decreased concentration of tartrate-resistant acid phosphatase, the drug constructs a positive anabolic window during the primary callus formation period of fracture healing, where the rate of bone formation is higher than the rate of bone resorption, and enhances the biomechanical fracture load of the new callus.

[0006] Preferably, the drug administration time window is limited to the period from hematoma organization to callus remodeling after fracture reduction and fixation; during this period, the drug uses a bidirectional decoupling mechanism to correct the pathological deviation of bone metabolism in osteoporosis, and improve the bone volume fraction and trabecular thickness parameters of newly formed trabecular bone at the fracture ends.

[0007] Preferably, the application includes establishing a drug efficacy evaluation procedure based on serum biochemical indicators; the procedure uses the increasing trend of serum type I procollagen N-terminal propeptide concentration and the decreasing trend of serum anti-tartrate acid phosphatase concentration as objective indicators of the initiation of a bidirectional decoupling mechanism; the drug preparation process is controlled to ensure that the active component can induce the objective indicators to appear during the callus formation period after administration.

[0008] Preferably, the application involves modulating osteoprotegerin and nuclear factor at the fracture site. The molar concentration ratio of B receptor activator ligands was used to inhibit secondary callus resorption; the unit dose of the drug was set sufficient to maintain osteoprotegerin in the fracture microenvironment at a saturated level binding to free nuclear factor. The concentration level of B receptor activator ligand prevents osteoclasts from being recruited during the callus sclerosis stage.

[0009] Preferably, the application utilizes the sustained expression of the Runx2 transcription factor to upregulate the synthesis levels of osteocalcin and bone alkaline phosphatase, promoting the deposition of calcium and phosphorus ions on the type I collagen fiber scaffold; the deposition process eliminates mineralization defects commonly found in the healing of osteoporotic fractures and increases the bone mineral density of newly formed callus.

[0010] Preferably, the drug is prepared as an oral formulation and maintains its ability to continuously inhibit intracellular glycogen synthase kinase-3 in osteoblasts throughout the fracture healing period. The active blood concentration of the drug; the inhibitory effect occurs simultaneously in bone marrow mesenchymal stem cells, inducing stem cells to differentiate into osteoblasts and inhibiting their differentiation into adipose cells.

[0011] Preferably, the application enhances the torsional strength of the fracture site through a bidirectional decoupling mechanism; the drug maintains the integrity of the bone collagen fiber network by inhibiting the degradation of type I collagen cross-linked C-terminal peptides, thereby enhancing the energy absorption capacity of the callus tissue under external loads.

[0012] Preferably, the positive anabolism window is calculated by the anabolism index. Perform quantitative characterization: ,in, The serum N-terminal propeptide concentration of type I procollagen at a specific time point t after drug intervention. This represents the baseline concentration before drug intervention. The serum antitartaric acid phosphatase concentration at a specific time point t during drug intervention. The baseline concentration before drug intervention was used; the effective dose of the drug was set to reduce the anabolic index. During the initial callus formation phase of fracture healing, the value remained in the positive range greater than 0.5.

[0013] Preferably, the application also includes the combined use of the bone-strengthening and blood-nourishing oral liquid with calcium and vitamin D preparations that provide substrates for bone matrix mineralization; in the combined system, the bone-strengthening and blood-nourishing oral liquid is responsible for reshaping the cellular balance of bone metabolism, while the calcium and vitamin D preparations are responsible for providing the material basis required for mineralization.

[0014] Preferably, the application is targeted at postmenopausal women with osteoporotic fractures who are in a state of estrogen withdrawal; the drug utilizes the Wnt signaling complex activation pathway, which is independent of estrogen receptors, to alternatively exert the biological effects of promoting osteogenic formation and inhibiting osteoclastization, thereby avoiding delayed fracture healing caused by low endogenous estrogen levels.

[0015] Compared with the prior art, the beneficial effects of the present invention are: 1. In the preparation of a drug for treating osteoporotic fractures, the Qiang Gu Sheng Xue oral liquid establishes a unique metabolic pattern in the osteoporotic fracture healing process that differs from conventional anti-osteoporosis treatments. Existing technologies, which simply inhibit bone resorption, typically reduce the rate of bone formation secondary to intercellular coupling signals, leading to a low-turnover bone state. This invention utilizes the synergistic effect of drug components to activate Wnt / The α-catenin signaling pathway directly drives the expression of Runx2 transcription factor to increase the rate of type I collagen synthesis, and induces osteoblasts to secrete osteoprotein to competitively block the RANKL / RANK signaling axis to activate osteoclasts. This mechanism breaks the traditional osteogenic-osteoclast coupling dependence in the fracture end microenvironment, and constructs a positive anabolic metabolic window with high osteogenic activity and low osteoclast activity, achieving independent and rapid bone accumulation without sacrificing bone turnover activity.

[0016] 2. To address the problem of rapid degradation of primary callus due to excessive recruitment of osteoclasts in an osteoporotic environment, during the fracture healing and remodeling period, the expression levels of tartrate-resistant acid phosphatase and type I collagen crosslinking C-terminal peptide are downregulated. This specifically inhibits the pathological reabsorption process of newly formed trabecular bone, ensuring that the primary callus formed by endochondral osteoblasts maintains sufficient volume and density, and smoothly transforms into a mature bone plate with superior mechanical properties. From the tissue structure level, this avoids the risk of delayed fracture healing or periprosthetic refracture caused by premature degradation of the bone matrix, and improves the biomechanical load-bearing capacity of the bone after healing.

[0017] 3. Provide a biological regulation program to correct pathological deviations in osteoporotic fractures. Targeting the physiological needs of different stages of fracture healing, it regulates calcium and phosphorus metabolism in the early stages of administration to provide a material basis for hematoma organization. During the critical callus formation and remodeling periods, it uses a bidirectional regulatory mechanism to widen the concentration gradient difference between bone formation markers and bone resorption markers, conforming to and strengthening the biochemical regulation of bone repair sequence. This corrects the imbalance between delayed collagen synthesis and accelerated matrix dissolution in osteoporotic patients, enabling the body's own bone homeostasis regulation network to be rapidly rebuilt at the site of injury, shortening the time required to reach clinical bone healing standards. Attached Figure Description

[0018] Figure 1 This is a flowchart illustrating the molecular regulatory pathway of the targeted bidirectional decoupling mechanism of the bone-strengthening and blood-generating oral liquid of the present invention. Figure 2 This is a schematic diagram showing the comparison of serum PINP and TRACP concentrations and the restoration of bone metabolism balance in each experimental group of this invention; Figure 3 This is a diagram illustrating the full-cycle application implementation architecture of the present invention based on active component quality control and dose closed-loop feedback. Detailed Implementation

[0019] The technical solution of the present invention is further explained so that those skilled in the art can more clearly understand the essence and features of the present invention. However, the following embodiments are only used to explain the present invention and do not constitute a limitation on the scope of protection of the present invention.

[0020] This invention proposes the application of a bone-strengthening and blood-nourishing oral liquid in the preparation of drugs for treating osteoporotic fractures. Utilizing the active components in the oral liquid, it activates the Wnt signaling complex within osteoblasts, constructing a specific temporal and spatial separation pattern between osteogenic and osteoclastic activities. This establishes a positive anabolic advantage during the healing window of osteoporotic fractures. The technical implementation of this scheme mainly revolves around three core dimensions: the molecular targeting effect of the active drug components, the pathophysiological matching of drug administration timing, and dynamic monitoring of efficacy based on serum biochemical indicators. The technical logic runs from top to bottom through the entire biological process from molecular signal transduction to the improvement of macroscopic bone biomechanical properties. This approach is applicable to the clinical treatment of osteoporotic fractures. In this context, there are unresolved objective obstacles at the application level. Conventional anti-osteoporosis drugs cannot effectively break the physiological osteogenic-osteoclast coupling dependence. In the osteoporotic state, this coupling mechanism is often in a pathological negative balance. Simply inhibiting osteoclasts reduces bone loss, but it also leads to a secondary decrease in osteogenic activity due to the interruption of coupling signals, resulting in low-turnover frozen bone. This is extremely detrimental to the fracture healing process, which requires rapid callus formation and remodeling. On the other hand, simple osteopromoting drugs may induce a synchronous upregulation of osteoclast activity due to the coupling mechanism, leading to secondary callus resorption. The present invention is configured to utilize a specific molecular regulatory network to independently and specifically block the transmission of osteoclast signals while promoting osteogenic formation.

[0021] Therefore, the drug used in this invention utilizes the active components in the Strong Bone and Blood Promoting Oral Solution to target and bind to glycogen synthase kinase-3 in the cytoplasm of osteoblasts. (GSK-3) By inhibiting phosphorylation activity, it blocks The ubiquitination and degradation pathway of β-catenin proteins, this molecular event enables... β-catenin accumulates in large quantities in the cytoplasm and translocates into the nucleus, where it binds to T-cell kinase / lymphocyte enhancer factor (TCF / LEF) and initiates the gene expression of downstream key osteogenic transcription factors Runx2 and Osterix. Direct upregulation of Runx2 increases the rate of type I collagen synthesis in osteoblasts, manifested as a rapid increase in serum N-terminal propeptide (PINP) concentration of type I procollagen, thereby driving the rapid conversion of cartilaginous callus to bony callus. Simultaneously, this drug utilizes the paracrine effect of osteoblasts responding to activation of the Wnt signaling complex to upregulate the synthesis and secretion of osteoprotegerin (OPG), a nuclear factor... RANKL, a soluble decoy receptor for beta-receptor activator, competitively binds to RANKL in the extracellular microenvironment at high concentrations of OPG, blocking the binding of RANKL to RANK receptors on the surface of osteoclast precursors, thereby inhibiting intracellular NF-κB. Activation of the B and c-Fos / NFATc1 signaling pathways leads to a decrease in the expression levels of anti-tartrate acid phosphatase (TRACP) and type I collagen cross-linked C-terminal peptide (S-CTX). Through the above mechanism, this invention successfully constructs a positive anabolic window in the primary callus formation period of fracture healing, where the rate of osteogenic formation is higher than the rate of bone resorption, thereby achieving rapid accumulation of bone mass and effective repair of bone microstructure. This solves the clinical problems of poor callus quality, easy secondary resorption, and refracture in osteoporotic environments.

[0022] In a specific implementation scenario of this invention, for postmenopausal patients with osteoporotic fractures, the application strictly limits the drug administration window to the period from hematoma organization to callus remodeling after fracture reduction and fixation. During this period, the drug utilizes the Wnt signaling activation pathway, which is independent of estrogen receptors, to alternatively exert an osteogenic effect, overcoming the delayed fracture healing caused by low endogenous estrogen levels. To ensure the controllability and reproducibility of the treatment effect, this invention establishes a pharmacodynamic evaluation procedure based on serum biochemical indicators. The increasing trend of PINP concentration and the decreasing trend of TRACP concentration in serum are defined as objective indicators of successful initiation of the bidirectional decoupling mechanism. This evaluation procedure requires the collection of venous blood samples at weeks 2, 4, and 8 after drug administration, and the above indicators are quantitatively detected using enzyme-linked immunosorbent assay (ELISA), and the anabolic index is calculated. The index is defined as follows: ,in, and The concentration at treatment time point t. and The baseline concentration is... If the value remains positive (greater than 0.5) during the primary callus formation period, it is determined that the positive anabolic window induced by the drug has been successfully established. This quantitative indicator not only verifies the biological activity of the drug, but also provides an objective basis for adjusting the dosing regimen in clinical practice.

[0023] Furthermore, to further inhibit secondary callus resorption, this invention regulates osteoprotegerin and nuclear factor at the fracture site. The molar concentration ratio of beta-receptor activator ligands is used to achieve this. The unit dose of the drug is set to be sufficient to maintain the concentration level of OPG in the local fracture microenvironment at a saturated level binding to free RANKL. In the specific preparation process, by controlling the extraction and concentration parameters, it is ensured that each oral liquid contains sufficient amounts of key components such as active astragaloside A and collagen peptides, so that after oral absorption, it can maintain sustained inhibition of intracellular GSK-3 in osteoblasts. The effective blood concentration of active drug, this sustained inhibitory effect not only occurs in mature osteoblasts, but also acts simultaneously on bone marrow mesenchymal stem cells, inducing differentiation into the osteogenic lineage and inhibiting differentiation into the adipose lineage, thereby increasing the number of osteoblasts at the cellular level, further consolidating the cellular basis of fracture healing. Ultimately, this precise regulation based on molecular mechanisms manifests as an increase in the torsional strength of the fracture site, enhanced energy absorption capacity of callus tissue under external loads, and prevention of refracture.

[0024] Example 1: This example describes the specific application of Qiang Gu Sheng Xue oral liquid in the preparation of drugs for treating osteoporotic fractures, specifically targeting the clinical situation of postmenopausal osteoporotic fractures accompanied by both high osteoclast activity and low osteogenic activity. In this scenario, due to low endogenous estrogen levels, the fracture microenvironment faces osteoblast inertia and osteoclast overactivity caused by an imbalance in the RANKL / OPG ratio, leading to delayed callus formation and an increased risk of secondary bone resorption. To address this situation, the active components of the drug are absorbed orally and distributed to the fracture site. They then enter the osteoblast cytoplasm via transmembrane transport, specifically binding to and inhibiting glycogen synthase kinase-3. (GSK-3) The inhibition of phosphorylation activity of ) blocks the phosphorylation activity of ) The ubiquitination and degradation cascade of β-catenin proteins promotes the degradation of undegraded proteins. β-catenin accumulates in the cytoplasm and translocates into the nucleus. After entering the nucleus... -catenin forms a transcriptional activation complex with T cell kinase / lymphocyte enhancer factor (TCF / LEF), which initiates the sustained expression of the downstream osteogenic transcription factor Runx2. Upregulation of Runx2 levels relieves the inhibitory state of osteoblast synthesis, drives the synthesis and secretion of type I procollagen N-terminal propeptide (PINP), and provides a matrix basis for cartilage callus mineralization.

[0025] Simultaneously, the drug utilizes osteoblasts as signal transduction hubs, activating the Wnt signaling pathway while inducing osteoblasts to increase the synthesis and paracrine release of osteoprotegerin (OPG). The high concentration of OPG released into the extracellular microenvironment acts as a soluble decoy receptor, competitively binding to free nuclear factor. Receptor activator ligand (RANKL) blocks the binding of RANKL to RANK receptors on the surface of osteoclast precursors. This blocking effect severs key upstream signals required for osteoclast differentiation and inhibits intracellular NF-κB. Activation of the B signaling pathway reduces the expression levels of anti-tartrate acid phosphatase (TRACP) and type I collagen cross-linked C-terminal peptide (S-CTX). Through this mechanism, this technique achieves uncoupling regulation of osteogenic promotion and osteoclast inhibition within a single therapeutic framework, resulting in atypical kinetic characteristics in serum biochemical assays: increased PINP concentration and decreased TRACP concentration. The calculated anabolic index... During the initial callus formation period, the value is maintained in a positive range greater than 0.5. This positive anabolic window ensures that the new callus is protected from pathological degradation by osteoclasts during its formation, allowing the volumetric density and connectivity of the trabecular bone at the fracture ends to recover during the callus remodeling period, thereby enhancing the biomechanical strength of the bone structure after healing.

[0026] Example 2: This example verifies the pharmacodynamic characteristics of Qiang Gu Sheng Xue oral liquid in a simulated postmenopausal osteoporotic fracture (OPF) biological environment and its ability to regulate the bone metabolic microenvironment. 120 SPF-grade female SD rats were used as bioreactors. A composite pathological model was constructed using bilateral ovariectomy (OVX) combined with femoral osteotomy to simulate fragility fractures under estrogen withdrawal. To eliminate individual differences in basal metabolic rate and background noise interference from surgical stress, a stratified randomized grouping strategy was adopted, with a sham-operated group serving as a physiological baseline reference. The experiment included a model control group (administered pure water to simulate the natural disease course), a positive control group (administered Xianling Gubao capsules at a dose of 0.27 g / kg, serving as an industry benchmark), and the present invention sample group (administered Qiang Gu Sheng Xue oral liquid at a dose of 10.0 mL / kg, i.e., four times the clinically equivalent dose, to verify the maximum efficacy boundary at the saturation dose). The administration period was 8 weeks, covering the complete healing cycle from hematoma organization to callus formation.

[0027] The experiment focused on monitoring the dynamic evolution of serum bone turnover markers to quantitatively assess the initiation of the bidirectional uncoupling mechanism. To address fluctuations in endogenous enzyme activity and measurement system errors in biological samples, data acquisition employed a double-antibody sandwich enzyme-linked immunosorbent assay (ELISA), with three repeated measurements at each time point to average the results and smooth noise. At week 8 of the drug intervention, the serum biochemical indicators in the experimental group exhibited a scissor-like effect. The concentration of type I procollagen N-terminal propeptide (PINP), a key indicator of bone formation, was significantly lower in the model control group due to the lack of estrogen stimulation. Maintaining a low level (28.97±5.26 ng / mL), the concentration was increased to 52.22±12.63 ng / mL, recovering to a level close to that of the sham-operated group. At the same time, the concentration of tartrate-resistant acid phosphatase (TRACP), a key indicator of bone resorption, was reduced from a pathologically high level (67.87±8.42 ng / mL) in the model control group to 52.69±12.29 ng / mL in the sample group of this invention. See Table 1, which summarizes the measured data of key biochemical indicators and calculated anabolic indices of each group at the end of week 8.

[0028] Table 1: Comparison of Bone Metabolic Biochemical Indicators and Biomechanical Performance among Experimental Groups in Week 8 Objective analysis based on the data in Table 1 shows that the sample group of this invention constructs a positive anabolic window by increasing PINP and simultaneously decreasing TRACP. >0.5), this biochemical regulation translates into improved physical properties of macroscopic tissue structures. In micro-computed tomography (Micro-CT) and three-point bending mechanical tests, the bone mineral density (BMD) of the sample group of this invention increased to 0.60±0.07 g / cm³, higher than the 0.52±0.05 g / cm³ of the model control group; and the maximum fracture load of the femur reached 140.06±17.42 N, indicating that the newly formed callus achieved substantial repair in terms of structural density and resistance to mechanical damage. The above data chain confirms that Qiang Gu Sheng Xue oral liquid can effectively correct the negative balance of bone metabolism in osteoporosis, without introducing additional bone growth factors, by regulating endogenous Wnt / Balancing the α-catenin and OPG / RANKL signaling pathways can improve the quality of fracture healing.

[0029] Example 3: This example combines Figures 1 to 3 Instructions for the application of Qiang Gu Sheng Xue oral liquid in the preparation of drugs for treating osteoporotic fractures, such as... Figure 1 As shown, the process begins with the intervention of a bone-strengthening and blood-nourishing oral liquid as a drug. This drug contains active components such as astragaloside A and collagen peptides, and its mechanism of action is to target and bind to GSK-3 within osteoblasts. To inhibit phosphorylation activity and block The degradation of catenin produces two parallel branches of biological effects, one of which is... -catenin enters the nucleus and binds to TCF / LEF to initiate Runx2 transcription factor expression, promoting the synthesis and increasing the synthesis rate of type I procollagen N-terminal propeptide, manifested as an increase in serum PINP concentration. Secondly, it utilizes the paracrine effect induced by Wnt signaling to upregulate osteoprotegerin OPG secretion, and inhibits osteoclast NF-κB by competitively binding and blocking RANKL / RANK binding. Activation of the B pathway ultimately leads to a bidirectional decoupling mechanism, which is confirmed by objective indicators, namely serum biochemical markers, to assess the anabolic index. A value >0.5 indicates an increase in PINP and a decrease in TRACP, which establishes a positive anabolic window during the primary callus formation period, thereby improving the quality of fracture healing by controlling secondary callus absorption and enhancing biomechanical fracture load.

[0030] like Figure 2 As shown in the figure, the horizontal axis represents three groups: the model control group, the positive control group, and the sample group of this invention. The vertical axis indicates the concentrations of PINP and TRACP, respectively, both in ng / mL. The model control group exhibits a pathological state with low PINP and high TRACP concentrations, showing negative metabolic characteristics. In contrast, the data from the sample group of this invention shows that the PINP concentration has recovered to a level similar to that of TRACP, with both values ​​maintained within the range of 50 ng / mL to 60 ng / mL, reflecting the re-established balance between osteogenic and osteoclastic activities after drug intervention. Figure 3 As shown, the preparation and quality control center serves as the standardized drug production end, encompassing an active ingredient extraction unit to control temperature and time parameters to ensure the retention rate of astragaloside A and other active ingredients, a fingerprint spectrum dual-calibration module to ensure batch-to-batch uniformity of active substances, and intelligent correction functions for process parameters. Next is the delivery of finished drugs to the clinical treatment environment. This stage involves patient dosing and metabolic regulation, specifically implementing dosing time window management from the hematoma organization stage to the callus remodeling stage, targeting the fracture local microenvironment to construct an osteogenic advantage window, and personalized dosing using oral formulations combined with calcium supplements. Finally, through regular biological sample collection, the data enters the efficacy evaluation and feedback center. This data analysis and decision-making end includes a biochemical index detection module to measure type I procollagen precursor peptide and tartrate-resistant acid phosphatase, an anabolic index calculation module to quantify the concentration difference between osteogenic and osteoclast-reducing components, and a dose closed-loop feedback system. This system dynamically outputs dose adjustment optimization instructions based on index deviation and sends them back to the clinical treatment end.

[0031] Example 4: This example aims to address the potential mechanistic and parameter issues regarding the active components and their bidirectional decoupling mechanism in Qiang Gu Sheng Xue oral liquid that may have existed in the previous examples. It constructs an integrated targeted repair solution, further clarifying its pharmacodynamic material basis and action boundaries through systematic experimental design and data verification. This ensures the interpretability of the technical solution at the molecular level and its reproducibility at the engineering level. In the field of chemical materials, regarding the definition of the active pharmaceutical components, this example introduces a fingerprint-pharmacodynamic correlation analysis procedure. Ten batches of Qiang Gu Sheng Xue oral liquid samples from different production cycles were selected, and characteristic fingerprint spectra were established using high-performance liquid chromatography (HPLC). By comparing the chromatographic peak areas of each batch with the corresponding batches in the osteoblast model using GSK-3... Inhibition rate was used to screen key chromatographic peaks that showed a high positive correlation with efficacy. Mass spectrometry (MS) confirmed that the peak with a retention time of 15.2 minutes corresponded to astragaloside A. A dual-standardization quality control system was established, clearly defining that the content of astragaloside A in each milliliter of oral solution should not be less than 0.1 mg, and the total nitrogen content of the bone fluid extract should not be less than 25 mg, serving as the quantitative material benchmark for the active pharmaceutical ingredient. The standardized preparation procedure for the bone fluid, which serves as the core matrix of the drug, is as follows: Fresh, quarantine-qualified pig tibias are taken, residual muscle connective tissue is removed from the surface, and the bone fragments are mechanically crushed to a particle size of less than 2 cm. The bone fragments are mixed with purified water at a mass ratio of 1:4 and placed in a high-pressure extraction tank at 121... Extraction was performed at 0.12 MPa for 4 hours; the extract was filtered through a 200-mesh vibrating screen to remove bone residue, and the filtrate was concentrated to 60% under vacuum at -0.08 MPa. The fluid extract has a relative density of 1.15 to 1.20. The total nitrogen content of the fluid extract is not less than 2.5 g / 100 mL as determined by the Kjeldahl method, and the calcium content is not less than 0.6 g / 100 mL as determined by EDTA titration. This grade of bone fluid contains high abundance of small molecule collagen peptides and free amino acids, which directly provide the necessary substrate for the synthesis of type I procollagen driven by Runx2 transcription factor after Wnt pathway activation, and provide bio-derived calcium and phosphorus ions that match the mineralization rate of bone matrix.

[0032] In the synergistic mechanism of compound formulation, in addition to astragaloside A directly targeting and inhibiting GSK-3... The active ingredients, including Codonopsis pilosula, polysaccharides, Ganoderma lucidum triterpenes, and black fungus extract, constitute an immune microenvironment regulatory unit, downregulating local TNF-α at the fracture site. and the expression level of IL-6 pro-inflammatory factor, blocking the effect of inflammatory signals on intracellular Wnt / -Catenin pathway cross-inhibition maintains the net capacity of intracellular signal transduction channels. Jujube extract, rich in cyclic adenosine monophosphate, regulates the permeability of calcium ion channels on the osteoblast membrane surface, synergistically acting as a mineralization substrate in bone fluid to ensure sufficient hydroxyapatite deposition during the rapid growth phase of newly formed bone trabeculae. At the physical structural level, a bidirectional decoupling mechanism is implemented to construct a positive anabolic window. In the field of algorithmic logic, addressing the black box of drug dosage parameters in the bidirectional decoupling mechanism, this embodiment constructs a dose-response gradient verification model. Using rat bone marrow mesenchymal stem cells as the in vitro test subject, a culture system containing serum containing different concentrations of Qiang Gu Sheng Xue oral liquid (concentration gradient set at 0%, 5%, 10%, 20%, and 40%) was set up. Western blotting technology was used to quantitatively detect intracellular calcium ions in osteoblasts. The nuclear translocation level of β-catenin and the protein concentration of OPG in culture supernatant detected by ELISA showed that as the concentration of drug-containing serum increased, Both β-catenin nuclear translocation levels and OPG secretion showed a non-linear, dose-dependent increase; when the serum drug concentration reached 20%, -catenin nuclear translocation levels reached their peak, and OPG secretion entered a plateau phase. Further increasing the concentration to 40% did not improve the above indicators; instead, a slight decrease in cell viability was observed. The in vivo dosage corresponding to a 20% drug-containing serum concentration, namely the aforementioned 10.0 mL / kg, was determined, and the optimal working window for the bidirectional decoupling mechanism was activated, providing solid experimental evidence for clinical dosing regimens.

[0033] Example 5: In the pharmaceutical engineering and clinical deployment process of applying Qiang Gu Sheng Xue oral liquid to the preparation of drugs for treating osteoporotic fractures, this example discloses an offline calibration and data filling procedure for dual calibration of active components. This procedure establishes a standardized detection platform based on high-performance liquid chromatography-mass spectrometry (HPLC-MS) technology to perform qualitative and quantitative analysis of the active components in each batch of bone fluid, astragalus, codonopsis, and other raw materials. By collecting no less than 50 batches of raw material samples, a fingerprint database containing the retention time, characteristic peak area, and mass-to-charge ratio of each component is constructed. Using this database and multivariate statistical analysis methods such as principal component analysis (PCA), the fluctuation range of active component content and quality control indicators are determined. For the two core indicator components, astragaloside A and total nitrogen in bone fluid, a correlation with intracellular GSK-3 in osteoblasts is established. A mathematical model of the dose-effect relationship between inhibition rate and OPG secretion was used to output the minimum effective concentration thresholds of each indicator component in each milliliter of oral liquid through regression analysis, namely, astragaloside A ≥ 0.1 mg / mL and total nitrogen content ≥ 2.5 g / 100 mL. During the production process, if the detection value of a batch of semi-finished products is lower than this threshold, the model calculation results are used to correct it by adding high-purity extract or adjusting the concentration factor until the standard is met.

[0034] Furthermore, considering the differences among individual patients in fracture type, degree of osteoporosis, and basal metabolic rate, this embodiment discloses a pre-deployment calibration and dosing regimen fine-tuning procedure. This procedure requires that before clinical application, patients undergo comprehensive baseline testing of bone metabolism biochemical indicators, including serum PINP, TRACP, S-CTX, and bone mineral density (BMD). Based on the test results, a pre-set algorithm model is used to calculate the patient's initial anabolic index. ,like A value below the preset pathological threshold, such as -0.5, indicates a severe negative bone metabolic balance. In this case, the protocol recommends initiating an intensive dosing regimen, using a high dose (1.5 to 2 times the clinically equivalent dose) for the first two weeks of the first treatment cycle to rapidly increase blood drug concentration and forcibly activate the Wnt signaling pathway. The indicator should be retested every 2 to 4 weeks during treatment, based on... The dosage is dynamically adjusted based on real-time changes until the index stabilizes within the target range of greater than 0.5 and is maintained until fracture healing. This procedure achieves personalized adaptation of the treatment plan through standardized initial assessment and dynamic feedback mechanism.

[0035] Example 6: This example discloses a standardized engineering procedure for the calibration of key process parameters and closed-loop dosage control in the preparation of Qiang Gu Sheng Xue oral liquid as a drug for treating osteoporotic fractures. The aim is to avoid potential process fluctuations and individual differences in industrial production and clinical application through quantitative experimental design and feedback control logic. To address the fluctuation in the extraction efficiency of active components during industrial preparation, a process calibration procedure based on response surface methodology (RSM) is established, selecting the ethanol volume fraction... Extraction temperature and extraction time As a key process control variable, the overall extraction rate of the target active component (mainly astragaloside A) was used. Using the set value as the response value, a second-order polynomial regression model describing the nonlinear mapping relationship between process parameters and extraction rate was constructed by executing a Box-Behnken design (BBD) experiment. Based on the extreme value analysis of this model, the optimal response value was calculated. The theoretical optimal operating point for reaching the maximum value was determined, and after adjustments based on the thermal conductivity characteristics of the industrial reactor, the key process parameters on the production line were ultimately locked at [value missing]. , and We have established process control benchmarks aimed at maximizing the retention of thermosensitive active components, ensuring the uniformity and stability of the molecular targeting material basis in each batch of drugs from the engineering source.

[0036] To address the fluctuations in drug response caused by individual patient differences in clinical treatment, a system based on the anabolic index was constructed and applied. A closed-loop feedback dose regulation algorithm is proposed, which presets a target range that can maintain a positive anabolic window without inducing metabolic overload. , set as During the monitoring phase of treatment, the system calculates the current status based on real-time serum biochemical data. The value is compared with the target range. If The algorithm outputs a command to increase the daily dosage by a preset step size. To enhance the activation intensity of the Wnt signaling pathway; if The algorithm outputs instructions to maintain the current dose or decrease it by one step. This helps avoid the risk of bone tissue structure disorder caused by excessive osteogenic activity.

[0037] It will be apparent to those skilled in the art that the present invention is not limited to the details of the exemplary embodiments described above, and that the present invention can be implemented in other specific forms without departing from the spirit or essential characteristics of the present invention.

[0038] 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 it. 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 spirit and scope of the technical solutions of the present invention.

Claims

1. The use of a bone-strengthening and blood-producing oral liquid in the preparation of a drug for treating osteoporotic fractures, the bone-strengthening and blood-producing oral liquid being made of bone fluid, Astragalus, Codonopsis, Ganoderma, Chinese date and black fungus, characterized in that, The application is applied to the preparation of a medicine for regulating the bone metabolic microenvironment of a fracture end through a bidirectional decoupling mechanism, and the bidirectional decoupling mechanism realizes quantitative improvement of the quality of fracture healing through the following molecular biological path: The active ingredient in the medicine targets the Wnt signal transduction complex in osteoblasts, inhibits the phosphorylation activity of glycogen synthase kinase-3 , blocks the degradation of β-catenin protein and promotes its nuclear entry, combines with T cell factor and lymphoid enhancer factor to start the expression of Runx2 transcription factor, increases the synthesis rate of type I collagen N-terminal propeptide in serum as a key indicator of bone formation, and drives the transformation of cartilage callus into osseous callus; Meanwhile, the bidirectional decoupling mechanism utilizes the paracrine effect produced by osteoblasts in response to the activation of the Wnt signaling complex to upregulate the synthesis and secretion of osteoprotegerin, which competitively binds to the nuclear factor B receptor activator ligand, blocks the binding of ligand to the RANK receptor on the surface of osteoclast precursors, inhibits the activation of the NF- B signaling pathway, and reduces the expression level of tartrate-resistant acid phosphatase in serum as a key indicator of bone resorption; Based on the increase of the concentration of the N-terminal propeptide of type I procollagen and the decrease of the concentration of tartrate-resistant acid phosphatase, the medicine creates a positive synthetic window in which the bone formation rate is higher than the bone resorption rate during the initial callus formation period of fracture healing, and improves the biomechanical fracture load of the newly formed callus.

2. Use of the Qianggu Shengxue oral liquid according to claim 1 in the preparation of a drug for treating osteoporotic fractures, characterized in that, The application limits the administration time window of the medicine to the hematoma organization period to the callus remodeling period after fracture reduction and fixation; the medicine uses the bidirectional decoupling mechanism to correct the pathological deviation of bone metabolism under the condition of osteoporosis during this period, and improves the bone volume fraction and trabecular thickness parameters of the newly formed bone trabecula of the fracture end.

3. Use of the Qianggu Shengxue oral liquid according to claim 1 in the preparation of a drug for treating osteoporotic fractures, characterized in that, The application includes establishing a pharmacodynamic evaluation procedure based on serum biochemical indicators; the procedure takes the upward trend of the concentration of the N-terminal propeptide of type I procollagen in serum and the downward trend of the concentration of tartrate-resistant acid phosphatase in serum as objective indications of the activation of the bidirectional decoupling mechanism; and the preparation process of the medicine is controlled to ensure that the active ingredients can induce the appearance of the objective indications during the callus formation period after administration.

4. The use of the Qianggu Shengxue oral liquid according to claim 1 in the preparation of a drug for treating osteoporotic fractures, characterized in that, The application modulates osteoprotegerin and nuclear factor at the fracture site. The molar concentration ratio of B receptor activator ligands was used to inhibit secondary callus resorption; the unit dose of the drug was set sufficient to maintain osteoprotegerin in the fracture microenvironment at a saturated level binding to free nuclear factor. The concentration level of B receptor activator ligand prevents osteoclasts from being recruited during the callus sclerosis stage.

5. Use of the Qianggu Shengxue oral liquid according to claim 1 in the preparation of a drug for treating osteoporotic fractures, characterized in that, The application uses the sustained expression of Runx2 transcription factor to up-regulate the synthesis levels of osteocalcin and bone alkaline phosphatase, and promotes the deposition of calcium and phosphate ions on the type I collagen fiber scaffold; the deposition process eliminates the mineralization defects commonly seen in osteoporotic fracture healing, and improves the bone mineral density of the newly formed callus.

6. Use of the Qianggu Shengxue oral liquid according to claim 1 in the preparation of a drug for treating osteoporotic fractures, characterized in that, The drug is prepared as an oral formulation and maintained at a blood plasma concentration capable of sustained inhibition of glycogen synthase kinase-3 activity within osteoblasts throughout the fracture healing period; the inhibitory effect is synchronized with bone marrow mesenchymal stem cells, inducing differentiation of the stem cells to the osteogenic lineage and inhibiting their differentiation to the adipogenic lineage.

7. Use of the Qianggu Shengxue oral liquid according to claim 1 in the preparation of a drug for treating osteoporotic fractures, characterized in that, The application improves the torsional strength of the fracture site through the bidirectional decoupling mechanism; the medicine maintains the integrity of the bone collagen fiber network by inhibiting the degradation of the C-terminal peptide of type I collagen cross-linking, and improves the energy absorption capacity of the callus tissue when subjected to external load.

8. Use of the Qianggu Shengxue oral liquid according to claim 1 in the preparation of a drug for treating osteoporotic fractures, characterized in that, The anabolic window is calculated by the anabolic index Quantitative characterization is performed: wherein Ccollagen(t) is the serum concentration of N-terminal propeptide of type I collagen at a specific time point t of the drug intervention, Ccollagen(0) is the baseline concentration before the drug intervention, Catp(t) is the serum concentration of tartrate-resistant acid phosphatase at a specific time point t of the drug intervention, Catp(0) is the baseline concentration before the drug intervention, the effective dose of the drug is set such that the anabolic index is maintained in the positive interval greater than 0.5 during the primary callus formation phase of fracture healing.

9. Use of the Qianggu Shengxue oral liquid according to claim 1 in the preparation of a drug for treating osteoporotic fractures, characterized in that, The application also includes the combined use of Qianggu Shengxue Oral Liquid, calcium agents that provide bone matrix mineralization substrates, and vitamin D preparations; Qianggu Shengxue Oral Liquid is responsible for remodeling the cytological balance of bone metabolism in the combined system, and calcium agents and vitamin D preparations are responsible for providing the material basis required for mineralization.

10. Use of the Qianggu Shengxue oral liquid according to claim 1 in the preparation of a medicament for treating osteoporotic fractures, characterized in that, The application is applied to postmenopausal women with osteoporotic fractures in the estrogen withdrawal state; the medicine uses the Wnt signal transduction complex activation pathway that does not depend on estrogen receptors to alternatively exert biological effects of promoting osteogenesis and inhibiting osteoclasts.

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