Application of citral in inhibition of synovial fibroblast-like cell proliferation and inflammatory response
The local administration of citral agents inhibits the proliferation and inflammatory response of synovial fibroblast-like cells, and solves the problems of limited efficacy, large side effects, poor compliance and high cost of existing RA treatments, providing a safe and effective multi-target treatment plan.
Patent Information
- Application Number
- CN202510831952.6
- Authority / Receiving Office
- CN · China
- Patent Type
- Applications(China)
- Current Assignee / Owner
- Filing Date
- 2025-06-20
- Publication Date
- 2025-09-05
AI Technical Summary
The existing RA treatment drugs have limited efficacy, significant side effects, poor patient compliance and high treatment costs, making it difficult to effectively control the proliferation and inflammatory response of synovial fibroblastoid cells.
Citral is used as a natural terpene compound to prepare topical administration preparations such as sustained-release gels, which inhibit the proliferation of synovial fibroblast-like cells and reduce the secretion of inflammatory factors. The concentration range is 0.131-13.1μM and the action time is 72 hours. It is combined with existing anti-rheumatic drugs to enhance the therapeutic effect.
Significantly inhibit the proliferation of synovial fibroblastoid cells, reduce the secretion of inflammatory factors, reduce side effects, improve patient compliance, reduce treatment costs, and provide multi-target treatment methods to control the pathological progress of RA.
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Figure CN120585803A_ABST
Abstract
Description
Technical Field
[0001] The present invention relates to the field of medicine, and in particular to an application of citral in inhibiting synovial fibroblast proliferation and inflammatory response. Background Art
[0002] Rheumatoid arthritis (RA) is an autoimmune disease characterized by erosive joint inflammation and progressive joint destruction. RA can develop at any age, and disability rates increase significantly with disease duration. Disability rates for patients with a disease course of 1-5 years, 5-10 years, and 10-15 years are 18.6%, 43.5%, and 48.1%, respectively. Therefore, treatment of RA is crucial for improving patients' quality of life and reducing disability.
[0003] Currently, RA treatment primarily relies on disease-modifying antirheumatic drugs (DMARDs), nonsteroidal anti-inflammatory drugs (NSAIDs), and glucocorticoids. These medications can alleviate disease activity, joint inflammation, and pain to a certain extent, but they also come with numerous drawbacks and adverse reactions. Specifically, the shortcomings of existing technologies are primarily reflected in the following aspects:
[0004] Limited efficacy: Although existing drugs can alleviate some symptoms, they are still unable to effectively control the progression of RA, especially in regulating synovial tissue proliferation and inflammatory responses. After long-term use, some patients may develop drug tolerance, resulting in reduced efficacy.
[0005] Significant side effects: Long-term use of antirheumatic drugs and glucocorticoids can lead to a variety of health problems, including gastrointestinal discomfort, liver and kidney damage, and cardiovascular disease, significantly reducing patients' quality of life. These side effects not only limit the dosage and duration of treatment but also increase treatment risks and medical burdens for patients.
[0006] Poor patient compliance: Due to the side effects and limited efficacy of existing medications, patients often face drug dependence and relapse during treatment. When patients gradually reduce or withdraw from medication, they often face relapses, which necessitates increasing the dosage or changing the treatment plan, further increasing the financial burden and psychological pressure on patients. Furthermore, the differences in responses and tolerance among different patients also make personalized treatment challenging, requiring constant adjustments to medication regimens, increasing the complexity of treatment, and reducing patient compliance.
[0007] High treatment costs: The treatment costs of existing drugs are high, especially for some patients who require long-term treatment, which imposes a heavy economic burden. In addition, due to the side effects and efficacy limitations of the drugs, patients may need to seek medical treatment frequently and change treatment plans, further increasing the cost of treatment. In view of these shortcomings of the existing technology, it is particularly important to explore new treatment methods to supplement the shortcomings of existing drug therapies. In order to solve the above problems, the applicant proposed an application of citral in inhibiting the proliferation of synovial fibroblasts and inflammatory responses. Summary of the Invention
[0008] The purpose of the present invention is to provide an application of citral in inhibiting synovial fibroblast proliferation and inflammatory response, so as to solve the problems in the prior art.
[0009] To achieve the above-mentioned purpose, the present invention provides the following technical solution: an application of citral in inhibiting the proliferation of synovial fibroblasts and inflammatory response, characterized in that the citral is used to prepare a drug for inhibiting the proliferation of synovial fibroblasts and reducing the inflammatory response, the effective concentration range of the citral in the drug is 0.131-13.1 μM, and the action time is 72 hours. By inhibiting the abnormal proliferation of synovial fibroblasts and reducing the secretion of inflammatory factors, the pathological process of rheumatoid arthritis is regulated, and the patient's joint function and quality of life are improved.
[0010] Optionally, the citral is a natural terpenoid compound with the chemical name of 3,7-dimethyl-2,6-octadienal, which has a clear chemical structure and biological activity.
[0011] Optionally, the drug is a local administration preparation so as to act directly on the synovial tissue of the joint, increase the local drug concentration, and reduce systemic side effects.
[0012] Optionally, the topical drug delivery preparation is a sustained-release gel, which can continuously and stably release citral into the synovial tissue of the joint, thereby prolonging the drug's duration of action and improving patient compliance.
[0013] Optionally, the drug further comprises a pharmaceutically acceptable carrier, diluent or excipient to optimize the stability, solubility or bioavailability of the drug and ensure the effectiveness and safety of the drug.
[0014] Optionally, the drug is used to prepare a pharmaceutical composition for treating rheumatoid arthritis, which is used alone or in combination with existing anti-rheumatic drugs to enhance the therapeutic effect or reduce adverse reactions, thereby improving the patient's quality of life.
[0015] Optionally, the concentration and duration of action of the citral are adjusted according to the specific conditions of the patient to achieve optimal therapeutic effect and safety.
[0016] Optionally, the application of citral in medicine is not limited to the treatment of rheumatoid arthritis, but also extends to other diseases caused by abnormal proliferation of synovial fibroblasts and inflammatory response, such as osteoarthritis and gouty arthritis.
[0017] Optionally, the method for preparing the sustained-release gel includes mixing citral with a polymer material, and forming a gel structure with a sustained-release function by a physical or chemical method to ensure the stable release of citral.
[0018] Optionally, the ratio of citral to existing anti-rheumatic drugs in the pharmaceutical composition is optimized based on clinical research data to achieve optimal therapeutic synergistic effects and reduce drug dosage and side effects.
[0019] Beneficial effects: The therapeutic effect is significantly improved:
[0020] This study experimentally demonstrated that citral, within a specific concentration range (0.131-13.1 μM), has a dual regulatory effect on synovial fibroblasts (FSCs), effectively inhibiting the proliferation of FLS and reducing the secretion of inflammatory factors such as IL-6 and TNF-α. This discovery provides a new and effective treatment for rheumatoid arthritis (RA), potentially significantly improving treatment outcomes and controlling disease progression.
[0021] Significantly reduced side effects:
[0022] Compared to existing anti-rheumatic drugs, nonsteroidal anti-inflammatory drugs, and glucocorticoids, citral, a natural terpenoid compound, has significant anti-inflammatory and anti-proliferative biological activities with relatively low toxic side effects. Long-term use of the citral treatment provided by the present invention can significantly reduce health problems caused by drug side effects, such as gastrointestinal discomfort, liver and kidney damage, and cardiovascular disease, thereby improving patients' quality of life.
[0023] Improved patient compliance:
[0024] Because citral has minimal side effects and is derived from natural plants, it is highly accepted by patients and their compliance is improved accordingly. Furthermore, citral can be administered via topical sustained-release gels, further improving patient convenience and comfort, and helping patients adhere to treatment over the long term.
[0025] Multi-target effect is superior to single-target solution:
[0026] This invention not only regulates the proliferation of synovial fibroblasts but also simultaneously inhibits the secretion of inflammatory factors, achieving a multi-targeted approach. Compared to single-target approaches, this multi-targeted approach can more comprehensively control the pathological process of RA, improve treatment efficacy, and reduce disease recurrence.
[0027] Reduced costs and easy promotion:
[0028] Citral is a natural ingredient, the raw materials are easily available, and the preparation process is relatively simple, so the cost is low. This feature makes the treatment method provided by the present invention easier to promote and apply in clinical practice, especially for patients with limited economic conditions, and has greater practical value and social significance. BRIEF DESCRIPTION OF THE DRAWINGS
[0029] Figure 1 Figure 2 is the inhibitory effect of citral on FLS proliferation;
[0030] Figure 2 This is a diagram showing the regulation of citral on the secretion of inflammatory factors. DETAILED DESCRIPTION
[0031] The following describes preferred embodiments of the present invention with reference to the accompanying drawings to make its technical content clearer and easier to understand. The present invention can be embodied in many different forms, and the scope of protection of the present invention is not limited to the embodiments mentioned herein.
[0032] This invention aims to explore the application of citral, a transdermal moxibustion component, in regulating the proliferation of synovial fibroblast-like cells (FLS) and the secretion of inflammatory factors, providing new ideas and effective treatment options for rheumatoid arthritis (RA). The specific technical solution is as follows:
[0033] Experimental groups and intervention methods
[0034] Synovial fibroblasts (primarily isolated FLS from RA patients) were used as experimental subjects and divided into a citral-treated group and a control group. The citral-treated group was further divided into three different concentration groups: 0.131 μM, 1.31 μM, and 13.1 μM. The corresponding concentration of citral was added to the FLS culture medium to achieve a final concentration in the culture medium of the preset value. The action time of citral was set to 72 hours to ensure that citral had sufficient time to act on the FLS.
[0035] Detection methods and key parameters
[0036] The cell proliferation rate was detected using the EDU method. EDU (5-ethynyl-2'-deoxyuridine) is a thymidine analog that can replace thymidine in the replicating DNA molecule during the cell proliferation period. The DNA replication activity is detected based on the specific reaction between EDU and Apollo fluorescent dye, thereby accurately reflecting the cell proliferation. The inflammatory factor secretion level was detected using the ELISA method. ELISA (enzyme-linked immunosorbent assay) is a detection method based on antigen-antibody specific reaction. It has the advantages of high sensitivity and strong specificity, and can accurately determine the content of inflammatory factors in the sample. The effective concentration range of citral is 0.131-13.1 μM. This concentration range was determined based on the results of previous experiments and can significantly affect the proliferation of FLS and the secretion of inflammatory factors. The action time is 72 hours. This length of time can ensure that citral has a stable and significant effect on FLS.
[0037] Implementation steps of the technical solution
[0038] Primary isolated FLS from RA patients were inoculated in appropriate culture medium and cultured in a 37°C, 5% CO2 incubator until the cells reached an appropriate density and state. Based on the experimental group, the appropriate concentration of citral was added to the FLS culture medium in each group, and the culture flask was gently shaken to evenly distribute the citral. The treated FLS culture flasks were then incubated in a 37°C, 5% CO2 incubator for an additional 72 hours. At the end of the incubation period, the FLS culture medium from each group was collected and centrifuged, and the supernatant was used to assay inflammatory cytokine secretion by ELISA. FLS cells were also collected for assaying cell proliferation rate by EDU assay.
[0039] By treating FLS with citral, it is expected to significantly inhibit the proliferation of FLS and reduce the secretion level of its inflammatory factors (such as IL-6, TNF-α), thereby providing a new idea and effective treatment option for the treatment of RA. The present invention reveals for the first time the mechanism of action of the moxibustion transdermal ingredient citral in regulating the proliferation of FLS and the secretion of inflammatory factors, providing a new target for the treatment of RA. As a natural ingredient, citral has the advantages of small toxic and side effects, easy access to raw materials, and simple preparation process. It is expected to become a safe and effective alternative therapy for the treatment of RA. The present invention experimentally verified the dual regulatory effect of citral on FLS in the concentration range of 0.131-13.1 μM (i.e., inhibiting proliferation and reducing the secretion of inflammatory factors), providing a scientific basis for the clinical application of citral.
[0040] Example 1: Experimental study on the regulation of synovial fibroblast proliferation by citral
[0041] Experimental conditions:
[0042] Cell source: Primary isolated synovial fibroblasts (FLS) from patients with rheumatoid arthritis (RA).
[0043] Citral concentrations: 0.131 μM, 1.31 μM, 13.1 μM.
[0044] Intervention duration: 72 hours.
[0045] Detection method: EDU method was used to detect cell proliferation rate.
[0046] Steps:
[0047] Cell culture:
[0048] The primary isolated FLS from RA patients were inoculated into cell culture plates and cultured in a constant temperature incubator at 37°C and 5% CO2 until the cells reached an appropriate density and state.
[0049] Grouping and intervention:
[0050] FLS were randomly divided into four groups: 0.131 μM citral treatment group, 1.31 μM citral treatment group, 13.1 μM citral treatment group and control group (no citral added).
[0051] The corresponding concentration of citral was added to the culture medium of each treatment group, and the same amount of culture medium was added to the control group.
[0052] Each group of cells was placed in a constant temperature incubator at 37°C and 5% CO2 and cultured for 72 hours.
[0053] Cell proliferation assay:
[0054] The EDU assay was used to detect the cell proliferation rate in each group. Specific procedures were performed according to the instructions of the EDU kit, including cell fixation, EDU labeling, and fluorescent staining.
[0055] The proliferation of cells in each group was observed and counted under a fluorescence microscope, and the proliferation rate was calculated.
[0056] result:
[0057] The experimental results showed that compared with the control group, the FLS proliferation rates in the 0.131μM, 1.31μM, and 13.1μM citral treatment groups were significantly reduced (P<0.05). Among them, the proliferation rate in the 13.1μM citral treatment group was the lowest, indicating that citral has a significant inhibitory effect on FLS proliferation within the 0.131-13.1μM concentration range.
[0058] Example 2: Experiment on the regulation of inflammatory factors secretion by citral in synovial fibroblasts Experimental conditions:
[0059] Cell source: Same as Example 1, primary isolated FLS from RA patients.
[0060] Citral concentration: same as in Example 1, 0.131 μM, 1.31 μM, 13.1 μM.
[0061] Intervention time: same as in Example 1, 72 hours.
[0062] Detection method: ELISA method was used to detect the secretion level of inflammatory factors (IL-6, TNF-α).
[0063] Steps:
[0064] Cell culture and grouping:
[0065] The cell culture and grouping steps were the same as in Example 1.
[0066] Intervention and collection of supernatant:
[0067] The corresponding concentration of citral was added to the culture medium of each treatment group, and the same amount of culture medium was added to the control group.
[0068] Each group of cells was placed in a constant temperature incubator at 37°C and 5% CO2 and cultured for 72 hours.
[0069] After the culture was completed, the supernatant of each cell group was collected and stored in a -80°C refrigerator for later use.
[0070] Inflammatory factor detection:
[0071] ELISA kits were used to detect IL-6 and TNF-α secretion levels in the cell supernatants of each group. Specific procedures were performed according to the ELISA kit instructions, including standard dilution, sample addition, incubation, plate washing, color development, reaction termination, and reading.
[0072] The absorbance of each well was read on a microplate reader, and the concentrations of IL-6 and TNF-α in the cell supernatant of each group were calculated according to the standard curve.
[0073] result:
[0074] The experimental results showed that compared with the control group, the secretion levels of IL-6 and TNF-α in the FLS supernatant of the 0.131μM, 1.31μM, and 13.1μM citral treatment groups were significantly reduced (P<0.05). Among them, the 13.1μM citral treatment group had the lowest level of inflammatory factor secretion, indicating that citral has a significant inhibitory effect on the secretion of inflammatory factors such as IL-6 and TNF-α by FLS within the concentration range of 0.131-13.1μM.
[0075] The above shows and describes the basic principles and main features of the present invention and the advantages of the present invention. It is obvious to those skilled in the art that the present invention is not limited to the details of the above exemplary embodiments, and that the present invention can be implemented in other specific forms without departing from the spirit or essential characteristics of the present invention. Therefore, from all points of view, the embodiments should be regarded as illustrative and non-restrictive. The scope of the present invention is defined by the appended claims rather than the above description, and it is intended that all changes that fall within the meaning and range of equivalents of the claims are included in the present invention. Any reference signs in the claims should not be construed as limiting the claim to which they relate.
[0076] In addition, it should be understood that although this specification is described in terms of implementation methods, not every implementation method contains only one independent technical solution. This narrative method of the specification is only for the sake of clarity. Those skilled in the art should regard the specification as a whole. The technical solutions in each embodiment can also be appropriately combined to form other implementation methods that can be understood by those skilled in the art.
Claims
1. An application of citral in inhibiting synovial fibroblast proliferation and inflammatory response, characterized in that: The citral is used to prepare a drug for inhibiting the proliferation of synovial fibroblasts and reducing inflammatory responses. The effective concentration range of the citral in the drug is 0.131-13.1 μM, and the action time is 72 hours. By inhibiting the abnormal proliferation of synovial fibroblasts and reducing the secretion of inflammatory factors, the pathological process of rheumatoid arthritis is regulated, and the joint function and quality of life of patients are improved.
2. The use according to claim 1, characterized in that The citral is a natural terpenoid compound with the chemical name of 3,7-dimethyl-2,6-octadienal, and has a clear chemical structure and biological activity.
3. The use according to claim 1, characterized in that The drug is a local administration preparation so as to directly act on the joint synovial tissue, increase the local drug concentration and reduce systemic side effects.
4. The use according to claim 3, characterized in that The topical drug delivery preparation is a sustained-release gel that can continuously and stably release citral into joint synovial tissue, thereby prolonging the drug's duration of action and improving patient compliance.
5. The use according to claim 1, characterized in that The drug further comprises a pharmaceutically acceptable carrier, diluent or excipient to optimize the stability, solubility or bioavailability of the drug and ensure the effectiveness and safety of the drug.
6. The use according to claim 1, characterized in that The drug is used to prepare a pharmaceutical composition for treating rheumatoid arthritis. The composition is used alone or in combination with existing anti-rheumatic drugs to enhance therapeutic effects or reduce adverse reactions, thereby improving the quality of life of patients.
7. The use according to claim 1, characterized in that The concentration and duration of action of the citral are adjusted according to the specific conditions of the patient to achieve optimal therapeutic effect and safety.
8. The use according to claim 1, characterized in that The application of citral in medicine is not limited to the treatment of rheumatoid arthritis, but also extends to other diseases caused by abnormal proliferation and inflammatory response of synovial fibroblasts, such as osteoarthritis and gouty arthritis.
9. The use according to claim 4, characterized in that The preparation method of the sustained-release gel comprises mixing citral with a polymer material, and forming a gel structure with a sustained-release function by a physical or chemical method to ensure the stable release of citral.
10. The use according to claim 6, characterized in that The ratio of citral to existing anti-rheumatic drugs in the pharmaceutical composition is optimized based on clinical research data to achieve optimal therapeutic synergistic effects and reduce drug dosage and side effects.