CN115845005A - Application of traditional Chinese medicine composition in adjusting expression level of PKA (protein kinase A) pathway-related protein of jejunum and preparing medicine - Google Patents
Application of traditional Chinese medicine composition in adjusting expression level of PKA (protein kinase A) pathway-related protein of jejunum and preparing medicine Download PDFInfo
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Abstract
The invention relates to a medicine for regulating protein or gene expression level and its application. Use of granule of folium astragali and fructus crataegi for regulating protein or gene expression level, wherein the regulation comprises increasing protein expression level, increasing gene expression level or decreasing gene expression level; wherein increasing the protein expression level comprises increasing the expression level of at least one protein selected from the group consisting of Ghrelin protein, GHSR protein, VIP protein, VPAC2 protein, rhoA/ROCK pathway-related protein, and PKA pathway-related protein; the increasing of the gene expression level comprises increasing of the expression level of at least one gene of a Ghrelin gene, a GHSR gene, a VIP gene, a VPAC2 gene, and an NPY gene; the reducing the expression level of the gene comprises reducing the expression level of a gene of POMC. The invention provides a theoretical basis for the wider application of astragalus, dendrobium and hawthorn granules.
Description
Technical Field
The application relates to the field of traditional Chinese medicines, in particular to a traditional Chinese medicine composition for adjusting the expression level of PKA (protein kinase activity) pathway-related proteins in the jejunum and application of the traditional Chinese medicine composition in preparation of medicines.
Background
The granule is a traditional Chinese medicine compound prepared from eleven medicinal materials such as astragalus and Chinese yam, has the effects of strengthening spleen and stomach, tonifying qi and consolidating exterior, and promoting digestion and removing food stagnation, and is mainly used for treating anorexia and monophagia caused by qi deficiency of spleen and stomach and dysfunction of spleen in transportation in children clinically. However, the application range of the ancient classic formula of astragalus, dendrobium and hawthorn particles is limited due to the problems of unclear pharmacodynamic mechanism and the like.
The invention is therefore proposed.
Disclosure of Invention
The invention mainly aims to provide a traditional Chinese medicine composition (astragalus, dendrobium and hawthorn granules) for adjusting the expression level of PKA (protein related to the gastrointestinal tract) pathway and an application of the traditional Chinese medicine composition in preparing a medicine. The invention discovers that the astragalus, dendrobium and hawthorn granules can obviously regulate the expression level of the related protein of the PKA pathway of the jejunum, thereby providing possibility for wider application of the astragalus, dendrobium and hawthorn granules, not only limiting to the treatment of the currently found diseases such as anorexia caused by deficiency of qi in spleen and stomach and dysfunction of spleen in transportation, but also expanding to other diseases related to the expression level of the protein, even including industrial production of the related protein and improving the yield of the related protein.
In order to achieve the above object, the present invention provides the following technical solutions.
The invention provides a use of a traditional Chinese medicine composition for adjusting the expression level of PKA (protein kinase) pathway-associated proteins in the jejunum, wherein the traditional Chinese medicine composition is astragalus mongholicus, dendrobium officinale and hawthorn granules, and the adjustment comprises the increase or decrease of the expression level of at least one protein in the PKA pathway-associated proteins in the jejunum.
The PKA pathway related proteins in the jejunum comprise p-PKA/PKA, p-eNOS and p-CaM proteins.
The modulation includes increasing p-PKA/PKA, p-eNOS protein expression levels, and decreasing p-CaM protein expression levels.
The second aspect of the invention provides an application of a traditional Chinese medicine composition in preparing a medicine for treating related diseases caused by the change of the expression level of a PKA (protein kinase) pathway related protein in the jejunum, wherein the traditional Chinese medicine composition is dendrobium officinale granules, and the change of the expression level of the protein comprises the reduction or the improvement of the expression level of the PKA pathway related protein in the jejunum.
The jejunal PKA pathway related proteins include p-PKA/PKA, p-eNOS, p-CaM proteins.
In summary, compared with the prior art, the invention achieves the following technical effects:
found that the granules of dendrobium nobile lindl and hawthorn can obviously regulate the expression level of the related protein of the PKA pathway of the jejunum, thereby mastering the pharmacodynamic mechanism of the granules of dendrobium nobile lindl and hawthorn, widening the application range of the granules of dendrobium nobile lindl and hawthorn according to the mechanism, and even being applied to the generation of the related protein.
The foregoing description is only an overview of the technical solutions of the present application, and the present application can be implemented according to the content of the description in order to make the technical means of the present application more clearly understood, and the following detailed description of the present application is given in order to make the above and other objects, features, and advantages of the present application more clearly understandable.
Drawings
Various additional advantages and benefits will become apparent to those of ordinary skill in the art upon reading the following detailed description of the preferred embodiments. The drawings are only for purposes of illustrating the preferred embodiments and are not to be construed as limiting the application. Moreover, like reference numerals are used to refer to like elements throughout. In the drawings:
FIGS. 1 to 3 show the effect of the granules of astragalus, dendrobium and hawthorn on the expression of proteins associated with the PKA pathway in the jejunal tissue (
n＝5)；
Wherein, in figure 1, a is blank group, b is model group, c is positive group, d is low dose group, e is medium dose group, f is high dose group;
p < 0.01 in fig. 2 compared to blank;
in comparison with the model set in figure 3, # P＜0.05。
Detailed Description
Embodiments of the present invention will be described in detail below with reference to the accompanying drawings. The following examples are only used to illustrate the technical solutions of the present application more clearly, and therefore are only used as examples, and the protection scope of the present application is not limited thereby.
Unless defined otherwise, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this application belongs; the terminology used herein is for the purpose of describing particular embodiments only and is not intended to be limiting of the application; the terms "including" and "having," and any variations thereof, in the description and claims of this application and the description of the above figures are intended to cover non-exclusive inclusions.
Reference herein to "an embodiment" means that a particular feature, structure, or characteristic described in connection with the embodiment can be included in at least one embodiment of the application. The appearances of the phrase in various places in the specification are not necessarily all referring to the same embodiment, nor are separate or alternative embodiments mutually exclusive of other embodiments. It is explicitly and implicitly understood by one skilled in the art that the embodiments described herein can be combined with other embodiments.
The invention researches the influence of astragalus, dendrobium and hawthorn granules on the expression of related proteins and genes based on the following experimental process.
1. Experimental Material
1.1 Experimental reagents and drugs
Experimental drugs: granule of astragalus root, dendrobium stem and hawthorn (KHYFH);
blank control: blank particles;
positive drug: children Qixing tea granules.
BCA protein quantification kit.
1.2 Experimental animals
70 SD rats (SPF grade) 1W after weaning, 60 + -10 g in body weight, and half in sex.
1.3 Experimental conditions
Barrier environment laboratory, the SPF level goes on, chooses SD experiment rat for use, and free drinking water, sufficient food, room temperature: 20-24 ℃, humidity: 30-50%, ventilation volume: 10-20 times per hour, and the fluorescent lamp can be used for lighting alternately in bright and dark.
1.4 Experimental instruments
TGL18W desk-top high-speed refrigerated centrifuge, ABI 7500 type real-time fluorescence quantitative PCR instrument, analytikjena UVP Chemstthio multifunctional gel imaging instrument, axiosceope A1 pathological imaging system.
2 method of experiment
2.1 dose design: the rat dose of the astragalus-dendrobium-hawthorn granules is 5.4g/kg, the rat dose is set as a medium dose, the high dose is set as 10.8g/kg, the low dose is set as 2.7g/kg, and the seven-star tea is set as 1.89g/kg, and the granules are prepared into the required concentration by using distilled water when in use. The blank granule dose was formulated according to the medium dose group at 5.4 g/kg. The above granules are prepared into desired concentration with distilled water.
2.2 animal groups: after SD rats are adaptively fed for 7 days, the SD rats are randomly divided into 10 blank control groups and 50 model groups by a random number table method; after the model is successfully prepared, the modeling components are randomly divided into 5 groups, namely a model control group (blank preparation particles), a positive control group (children's Qixing tea particles), a high, medium and low dose group of Qihuzha particles, and 10 groups of each group.
2.3 model preparation: modeling by adopting a etiology simulation method. The special molding feed consists of fish floss, corn flour, milk powder, soybean flour, white sugar, fresh eggs, fresh fat meat and other high-fat high-protein high-sugar components. The blank control group is fed with conventional feed, the modeling group is fed with special feed, and the feeding period is 28 days with sufficient food and free water. The rats were monitored daily for food intake and weight change, and the criteria for successful model preparation were: the food intake of the rats in the model group is reduced by 40-60% averagely compared with that in the blank control group, and/or the body weight of the rats is 10-15% lower than that in the blank control group.
2.4 dosage and method of administration: 7 days after the model building is successful, the rats of each group are fed with the conventional feed, and the administration is carried out by respectively gavage the stomach of each group according to the designed dose, once a day and continuously for 42 days. The normal group and model animals were given equal amounts of distilled water. After dosing, animals were sacrificed by anesthesia and samples were collected. Taking antrum and empty tissue, fixing a part of formaldehyde, and storing a part of formaldehyde at-80 deg.C.
3. Detecting the index
3.1 food consumption monitoring
From the start of molding to the end of dosing, rats were dosed 2 times per week and their feed intake was recorded, and the change in food intake was recorded for each group.
3.2 weight monitoring
From the start of the molding to the end of the administration, the body weight of the rats was measured 2 times and recorded, and the change in body weight of each group of rats was recorded.
3.3 detection of VPAC signal path related protein: western Blot assay PKA, p-PKA, eNOS, p-eNOS, caM, p-CaM protein expression.
3.4 statistical treatment
Statistical processing was performed using the SPSS 16.0 software system, and the measured data were expressed as mean. + -. S. The comparison between groups was performed by one-way ANOVA, by means of the test of variance, t test was performed when the variance was uniform, dunnett's T test was performed when the variance was irregular, and P < 0.05 indicated that the difference was statistically significant.
4 results of the experiment
4.1 evaluation of the model
4.1.1 general observations: during the molding and administration period, the blank animals had normal activity, glossy hair, no abnormality in diet and urination and defecation, and good spirit. During the modeling period, after one week of feeding the special feed, the animals begin to become thin, the hair is not glossy, the activity is basically normal, the stool becomes thin and loose, and the above situation is relieved after 3 weeks of modeling, which may be related to the initial adaptation of the animals to the special feed.
4.1.2 animal food intake: the blank group is prepared by taking 10 animals, wherein the animals are half male and half female, and the animals are bred in cages, 2 cages and 5 animals in each cage. The model building group actually takes 60 animals, each half of the animals are raised in cages, and the animals are divided into 12 cages, and each cage is provided with 5 animals. Compared with the blank group, the food intake of the animals is obviously reduced after one week of molding, and the food intake lasts for 4 weeks after molding. The food intake is reduced by 47.5%. According to the standard, the molding is successful (the food intake is reduced by 40-60%).
4.1.3 animal weights: compared with a blank group, the animal body weight is obviously reduced after the model is made for 1 week, and the difference is significant and lasts until the model is made for 3 weeks; the animals lost weight at week 4 of molding and lost 8.6% of weight, but there was no significant difference compared to the blank injection, and it may be related to the animals' adaptation to the special feed and increased food intake.
4.2 Effect of granule containing astragalus, dendrobium and hawthorn on animal food intake
After the model is made, 50 animals are randomly selected and divided into 5 groups, wherein the model group, the positive group, the astragalus mongholicus, dendrobium and hawthorn particle size are high (10.8 g/kg), medium (5.4 g/kg) and low (2.7 g/kg), 10 animals are fed in each group, the animals are male and female, and each group is divided into 2 cages to calculate the food intake. Each group was gavaged with the corresponding drug. Before administration, the food intake of the model animals was significantly reduced (by 45.81%) compared to the blank group, and by the end of the administration, food intake was restored, but still reduced by 32.46% compared to the blank group. Compared with the model group, the food intake of animals in the high and medium dose group with astragalus, dendrobium and hawthorn granules is obviously increased after 3 weeks of administration, and after the administration is finished, the food intake is increased by 43.58 percent and 39.10 percent respectively compared with the model group, the effect of the low dose group is relatively weaker, and the food intake is increased by 19.23 percent compared with the model group. The food intake of the positive group is increased by 41.67 percent compared with that of the model group. The results are shown in Table 1.
TABLE 1 influence of granule of astragalus, dendrobium and hawthorn on food intake of animals (
n＝10)
4.3 Effect of granule containing Astragalus Membranaeus and Crataegus pinnatifida on animal weight
Compared with the blank group, the model group animals grew slowly, and by the end of the administration, the weight lost 11.83%. Compared with the model group, the high and medium dose group containing astragalus, dendrobium and hawthorn granules can obviously promote the weight increase of animals, after the administration is finished, the weight is respectively increased by 13.42 percent and 11.14 percent, and the low dose group has relatively weak effect and the weight is increased by 4.82 percent. The results are shown in Table 2.
TABLE 2 influence of granule of Astragalus, dendrobii and Crataegus pinnatifida on animal body weight: (
n＝10)
4.4 influence of granule containing astragalus, dendrobium and hawthorn on expression of PKA pathway-associated protein in jejunum
Compared with the blank group, the p-PKA/PKA and p-eNOS protein expressions in the jejunum tissues of the model rats are obviously reduced, the p-CaM protein expression is obviously increased, and the difference is significant. Compared with the model group, the high and medium dose of the astragalus-dendrobium-hawthorn granules can obviously increase the expression levels of p-PKAPKA and p-eNOS proteins in jejunum tissues and can obviously reduce the expression of p-CaM proteins, and the difference is significant. The results are shown in FIGS. 1 to 3.
5. Conclusion
5.1 Effect of granule of Astragalus Membranaeus and Crataegus pinnatifida on expression of PKA pathway-associated protein in jejunum
Both p-PKA and p-eNOS are downstream signaling proteins of VAPC2, and up-regulation or down-regulation of their expression levels can be used as evidence of VIP/VPAC2 signaling pathway activation or inhibition. Activation of Protein Kinase A (PKA) following VIP activation in the jejunum causes smooth muscle cell hyperpolarization, ca 2+ Decreased inflow, decreased CaM activity (decreased expression), smooth muscle relaxation; in addition, VIP is combined with receptor to generate cascade amplification effect, which can stimulate endothelial type NOS (eNOS) to increase NO generation, induce relaxation of smooth muscle, and promote relaxation of jejunum. The experimental result shows that compared with a blank group, the p-PKA/PKA and p-eNOS protein expressions in the jejunum tissue of a model rat are obviously reduced, the p-CaM protein expression is obviously increased, and the difference is significant. Compared with the model group, the high and medium dose of the astragalus-dendrobium-hawthorn granules can obviously increase the expression levels of p-PKAPKA and p-eNOS proteins in jejunum tissues and can obviously reduce the expression of p-CaM proteins, and the difference is significant.
Finally, it should be noted that: the above embodiments are only used for illustrating the technical solutions of the present application, and not for limiting the same; although the present application has been described in detail with reference to the foregoing embodiments, it should be understood by those of ordinary skill in the art that: the technical solutions described in the foregoing embodiments may still be modified, or some or all of the technical features may be equivalently replaced; such modifications and substitutions do not depart from the spirit and scope of the present disclosure, and the present disclosure should be construed as being covered by the claims and the specification. In particular, the technical features mentioned in the embodiments can be combined in any way as long as there is no structural conflict. The present application is not intended to be limited to the particular embodiments disclosed herein but is to cover all embodiments that may fall within the scope of the appended claims.
Claims (5)
1. The application of a traditional Chinese medicine composition in regulating the expression level of PKA (protein kinase activity) related proteins in the jejunum is characterized in that the traditional Chinese medicine composition is astragalus, dendrobe and hawthorn granules, and the regulation comprises the step of increasing or reducing the expression level of at least one protein in the PKA related proteins in the jejunum.
2. The use of a Chinese medicinal composition of claim 1 for modulating the expression level of a protein associated with the PKA pathway in the jejunum, wherein the protein associated with the PKA pathway in the jejunum comprises p-PKA/PKA, p-eNOS, p-CaM proteins.
3. The use of a Chinese medicinal composition for modulating the expression level of a PKA pathway related protein in the jejunal region according to claim 2, wherein said modulation comprises increasing the expression level of p-PKA/PKA, p-eNOS proteins, and decreasing the expression level of p-CaM proteins.
4. The application of a traditional Chinese medicine composition in preparing medicines for treating diseases caused by the change of the expression level of a PKA (protein kinase) pathway related protein in the jejunum is characterized in that the traditional Chinese medicine composition is astragalus, dendrobe and hawthorn particles, and the change of the expression level of the protein comprises the reduction or the increase of the expression level of the protein related to the PKA pathway in the jejunum.
5. The use of a Chinese medicinal composition according to claim 4 for the preparation of a medicament for treating diseases associated with altered expression levels of PKA pathway associated proteins in the jejunum, wherein said PKA pathway associated proteins comprise p-PKA/PKA, p-eNOS, p-CaM proteins.
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| CN1879821A (en) * | 2006-04-29 | 2006-12-20 | 贵州宏奇药业有限公司 | A method for preparing sugar-free granule with astragalus root, dendrobium stem and haw thorn fruit |
| CN110812455A (en) * | 2019-11-01 | 2020-02-21 | 葵花药业集团北京药物研究院有限公司 | A liquid oral preparation for treating children anorexia and monophagia, and its preparation method and application |
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|---|---|---|---|---|
| CN1879821A (en) * | 2006-04-29 | 2006-12-20 | 贵州宏奇药业有限公司 | A method for preparing sugar-free granule with astragalus root, dendrobium stem and haw thorn fruit |
| CN110812455A (en) * | 2019-11-01 | 2020-02-21 | 葵花药业集团北京药物研究院有限公司 | A liquid oral preparation for treating children anorexia and monophagia, and its preparation method and application |
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