Application of cyanidin-3-O-glucoside in preparation of functional food for preventing bad intergenerational heredity

Abstract

The invention discloses an application of cyanidin-3-O-glucoside in preparation of a functional food for preventing poor intergenerational heredity. Experimental results show that the cyanidin-3-O-glucoside can be used for regulating glycolipid metabolism disorder of the offspring, and can be used for improving the uterine hypertrophy of the female offspring. The intervention of the cyanidin-3-O-glucoside of the parent can realize regulation and control of glucose and lipid metabolism disorder by improving insulin resistance and non-alcoholic fatty liver of the offspring, and can realize improvement of female offspring uterine hypertrophy by relieving the uterus volume and cross sectional area increase of the offspring and excessive cell proliferation. According to the invention, a new choice is provided for the functional food for preventing poor intergenerational inheritance, and a new application is also provided for the cyanidin-3-O-glucoside.

Description

Technical Field

[0001] The present invention belongs to the field of biomedicine technology, and relates to the application of cyanidin-3-O-glucoside in the preparation of functional foods, and more specifically to the application of cyanidin-3-O-glucoside in the preparation of functional foods for preventing adverse intergenerational inheritance. Background Art

[0002] A high-fructose diet can lead to lipid metabolism disorders and is one of the main dietary patterns that cause obesity, hyperglycemia and hyperlipidemia. The health problems caused by a high-fructose diet can also seriously threaten the health of offspring. It is inherited to offspring through epigenetic modifications, affecting the offspring's lipid metabolism and reproductive health, manifesting as adverse intergenerational inheritance. Anthocyanins are one of the important sources of natural functional pigments in food. They are widely present in dark fruits and vegetables. They have multiple functions such as hypoglycemic, anti-oxidant, anti-inflammatory and epigenetic regulation. They are potential functional factors for improving lipid metabolism disorders and adverse intergenerational inheritance effects induced by high fat and high fructose.

[0003] Cyanidin-3-O-glucoside (C3G) is the most easily available and most abundant anthocyanin in nature. It has a good regulatory effect on metabolic disorders caused by a high-fat and high-sugar diet, but its protective effect against the adverse intergenerational inheritance caused by a high-fructose diet has not been reported. Summary of the invention

[0004] In view of the shortcomings of the prior art, the present invention provides the use of cyanidin-3-O-glucoside in the preparation of functional foods for preventing adverse intergenerational inheritance, which can produce a protective effect against adverse intergenerational inheritance caused by a high-fructose diet, and provide a scientific basis for functional products for preventing adverse intergenerational inheritance.

[0005] The first object of the present invention is to provide the use of cyanidin-3-O-glucoside in the preparation of functional foods for preventing adverse intergenerational inheritance.

[0006] According to further features of the application of the present invention, the adverse intergenerational inheritance is lipid metabolism disorder in the offspring or uterine hypertrophy in the female offspring.

[0007] Preferably, the offspring glucose and lipid metabolism disorder is selected from one of the following: glucose intolerance; insulin resistance; abnormal total cholesterol, total triglycerides, high-density lipoprotein cholesterol and low-density lipoprotein cholesterol in plasma; non-alcoholic fatty liver.

[0008] Preferably, the female offspring uterine hypertrophy is selected from one of the following: increased offspring uterine volume; increased uterine myometrium area; increased endometrial area; and excessive proliferation of offspring uterine cells.

[0009] The second object of the present invention is to provide a functional food for preventing adverse intergenerational inheritance.

[0010] The functional food of the present invention contains cyanidin-3-O-glucoside.

[0011] The functional food of the present invention further comprises acceptable auxiliary materials.

[0012] The functional food of the present invention can be prepared into one of the following types: tablets, powders, pills, capsules, granules, dripping pills, and oral liquids.

[0013] The present invention uses an appropriate dose of cyanidin-3-O-glucoside to study the protective effect on the intergenerational inheritance of glucose and lipid metabolism disorders and uterine hypertrophy in female offspring.

[0014] The experimental results show that the intake of cyanidin-3-O-glucoside by the parents can alleviate the adverse intergenerational genetic effects. The C3G diet of the parents can significantly improve the intergenerational inheritance of insulin resistance and non-alcoholic fatty liver disease. The C3G diet of the parents can also alleviate the increase in the uterine volume and uterine cross-sectional area of ​​the offspring, and alleviate the symptoms of uterine hypertrophy by alleviating the excessive proliferation of uterine myometrial cells. Therefore, the intake of cyanidin-3-O-glucoside by the parents can effectively protect the sugar and lipid metabolism function and uterine health of the offspring, providing a new choice for functional foods to prevent adverse intergenerational inheritance, and also providing a new application for this substance. BRIEF DESCRIPTION OF THE DRAWINGS

[0015] Figure 1 This is the animal experiment design in Examples 1 and 2.

[0016] Figure 2 The HFHF diet of the father generation induced the glucose metabolism disorder of the F1 offspring and the improvement effect of C3G in Example 1. Wherein, A is the initial weight of the F1 mice; BC is the final weight of the F1 mice; D is the fasting blood glucose of the F1 mice; EF is the OGTT test and the area under the curve of the F1 mice; GH is the ITT test and the area under the curve of the F1 mice.

[0017] Figure 3 The HFHF diet induced lipid metabolism disorders in F1 offspring and the improvement effect of C3G in Example 1. AD is the plasma TC, TG, HDL-C and LDL-C content; E is the hematoxylin-eosin stained section of the liver; F is the liver non-alcoholic fatty liver score.

[0018] Figure 4 The uterine structure changes of F1 mice in Example 2. Among them, A is the uterine morphology change; B is the uterine volume statistics; C is the uterine section; D is the uterine basement and endometrial area statistics.

[0019] Figure 5The distribution of PCNA expression in the uterus of F1:ND mice in Example 2. A is PCNA immunohistochemical staining, Endometrium stroma (ES), endometrial stroma; Luminal epithelium (LE), endometrial epithelium; Myometrium (M), uterine muscle layer; B is the proportion of PCNA positive cells and the proportion in each layer.

[0020] Figure 6 The PCNA expression distribution in the myometrium of F1:ND mice in Example 2. A is the fluorescence expression image; B is the statistical analysis of PCNA expression. DETAILED DESCRIPTION

[0021] 1. Preparation and purification of C3G

[0022] Cyanidin-3-O-glucoside (C3G) is extracted and purified from black bean husk and mulberry resources. The raw materials can be black bean husk and mulberry dried fruit (References: [1] Li Xusheng, Jiang Xinwei, Shan Xiaoyu, et al. Medium pressure liquid chromatography preparation and structural identification of black bean husk anthocyanins [J]. Food and Fermentation Industries, 2018, 44(06): 51-57. DOI: 10.13995 / j.cnki.11-1802 / ts.015859.; [2] Ran Guojing, Jiang Xinwei, Li Haoyi, et al. Rapid preparation of cyanidin-3-glucoside monomer from mulberry using medium pressure preparative liquid chromatography [J]. Food Science, 2019, 40(03): 94-100.). The purified C3G has a purity of 97% at 280nm and is used in the following experiments.

[0023] 2. Establishment of high-fat and high-fructose model mice

[0024] Twenty-eight 6-week-old C57 / BL6J male mice were purchased from Zhejiang Weitong Lihua Experimental Animal Technology Co., Ltd. Figure 1As shown, F0 mice were randomly divided into four groups (n=7) according to the weight stratification method: normal diet (ND) and normal saline (ND+Vehicle), high fat and high fructose (HFHF) diet and normal saline (HFHF+Vehicle), high fat and high fructose diet and C3G intervention (HFHF+C3G), high fat and high fructose diet and metformin (Met) intervention (HFHF+Met). During the 4-week modeling phase, ND+Vehicle was fed with normal diet (10 kcal% normal feed) and normal drinking water, and the other three groups were fed with high fat and high fructose diet (10% fructose water and 60 kcal% high fat feed). In the subsequent 8-week intervention phase, HFHF+C3G received C3G (100 mg / kg·bw) gavage intervention, HFHF+Met received Met (200 mg / kg·bw) gavage intervention, and ND+Vehicle and HFHF+Vehicle received an equal amount of normal saline by gavage.

[0025] 3. Grouping and treatment of F1 offspring animals

[0026] After the intervention, the sperm of F0 mice was fertilized with the eggs of wild-type female mice in vitro (IVF) to obtain F1 offspring. The F1 offspring mice with four different paternal dietary backgrounds were divided into ND (10kcal% ordinary feed) and high-fat diet (HFD, 60kcal% high-fat feed) after sexual maturity. Mice in each group were free to eat and drink water. All feed and drinking water were replaced every 7 days. After 8 weeks of feeding, they were euthanized and collected. The experimental animals were raised in the Animal Management Center of Jinan University, with a 12-hour light day and night alternation, a temperature maintained at 22-26°C, and a relative humidity maintained at 60-80%.

[0027] F1 mice were fasted for 12 hours before euthanasia, anesthetized with isoflurane after weighing, and the eyeballs were removed for blood collection; after cardiac perfusion, the liver, testis, uterus and other organs were removed and weighed and recorded. The liver was fixed in 4% paraformaldehyde, the left testis was fixed in testis fixative, and the left uterus was fixed in 4% paraformaldehyde for paraffin embedding and sectioning; the right testis and right uterus were quickly frozen in liquid nitrogen and stored at -80°C.

[0028] 4. Glucose tolerance test and insulin tolerance test

[0029] After six weeks of intervention, the mice were subjected to an oral glucose tolerance test (OGTT), and the treatment before the test was the same as before the fasting blood glucose test. After fasting for 6 hours, the mice were gavaged with a 20% glucose solution at a dose of 1.5 g / kg·bw, and the tail blood of the mice was collected at 0, 15, 30, 60 and 120 minutes after gavage, and the values ​​were measured and recorded using a blood glucose meter. After seven weeks of intervention, the mice were subjected to an insulin tolerance test (ITT), and the treatment before the test was the same as before the fasting blood glucose test. After fasting for 6 hours, the mice were intraperitoneally injected with insulin at a dose of 0.75 U / kg·bw, and the tail blood of the mice was collected at 0, 20, 40, 60 and 120 minutes after injection, and the values ​​were measured and recorded using a blood glucose meter.

[0030] 5. Determination of total cholesterol, total triglycerides, high-density lipoprotein and low-density lipoprotein

[0031] The determination methods of total cholesterol (TC), triglyceride (TG), low density lipoprotein cholesterol (LDL-C), high density lipoprotein cholesterol (HDL-C), aspartate aminotransferase (AST) and alanine aminotransferase (ALT) in plasma were all performed according to the corresponding kits of Nanjing Jiancheng Biotechnology Co., Ltd. The insulin level of mice was determined by ELISA kit (Beijing Qisong Biotechnology Co., Ltd.) according to the standard operating method. HOMA-IR and HOMA-β index were calculated according to formula 1-1 and formula 1-2.

[0032] HOMA-IR = (plasma insulin (mIU / L) × fasting blood glucose (mmol / L)) / 22.5 (1-1)

[0033] HOMA-β = (20×plasma insulin (mIU / L)) / (fasting blood glucose (mmol / L)-3) (1-2) 6. Hematoxylin-eosin staining

[0034] (1) The samples were fixed in 4% paraformaldehyde for more than 24 h for preparation of paraffin sections.

[0035] (2) Take 5-8 μm paraffin sections, heat them, and then stain them in hematoxylin solution for 3-5 minutes. Wash them with tap water, differentiate them with hematoxylin differentiation solution, wash them with water, reblue them with bluing solution, and rinse them with running water.

[0036] (3) Dehydrate the sections in 85% and 95% graded alcohol for 5 min each, and then stain them in eosin solution for 5 min.

[0037] (4) Gently immerse the sections in pure water twice, for 5 seconds and 10 seconds respectively, and examine the staining effect under a microscope.

[0038] (5) Seal the sections with glycerol-gelatin mounting medium and collect and analyze microscopic images.

[0039] 7. Oil red staining

[0040] (1) Immerse the paraffin sections in oil red staining solution, cover and stain in the dark for 8-10 minutes.

[0041] (2) Take out the slices, let them sit for 3 seconds, then dip them in 60% isopropanol twice, for 3 seconds and 5 seconds respectively. Take out the slices and dip them in pure water twice, for 10 seconds each time.

[0042] (3) Remove the slices, leave them for 3 seconds, then immerse them in hematoxylin for 3-5 minutes, and then rinse them in pure water three times for 5 seconds, 10 seconds, and 30 seconds each.

[0043] (4) Differentiate with hematoxylin solution (with 60% alcohol as solvent) for 2-8 seconds; then rinse twice with pure water, each time for 10 seconds.

[0044] (5) Bluing solution for 1 second. Then gently immerse the sections in pure water twice, for 5 seconds and 10 seconds respectively, and examine the staining effect under a microscope.

[0045] (6) Seal the sections with glycerol-gelatin mounting medium and collect and analyze microscopic images.

[0046] 8. Scoring of Non-alcoholic Fatty Liver Disease in Mice

[0047] With reference to the diagnostic guidelines for non-alcoholic fatty liver disease (2010 revised edition), the livers of mice in each group were scored for non-alcoholic fatty liver activity (NAFLD activity score, NAS). The specific scoring rules are shown in Table 1.

[0048] Table 1: Non-alcoholic fatty liver disease scoring criteria

[0049]

[0050] IX. Uterine morphology and section statistics

[0051] (1) Uterine morphology photography

[0052] Place the unilateral uterus in a studio and photograph the uterine morphology under a stable light source. Use Image J to measure the length and width of the unilateral uterus, and calculate the uterine volume by approximating the unilateral uterus as a cylinder. Uterine volume = length × π × width 2 .

[0053] (2) Uterine cross-sectional area statistics

[0054] Image J was used to perform statistical analysis on the 40× microscope images of the hematoxylin-eosin-stained sections of the uterus, and the areas of the myometrium and endometrium were selected and calculated.

[0055] 10. Immunohistochemistry

[0056] (1) Slice, bake, dewax and perform antigen repair. Place tissue sections in citric acid antigen repair buffer (pH 6.0) and perform antigen repair in a microwave oven. Boil the repair solution on medium heat for 8 minutes, turn off the heat for 8 minutes to keep warm, and then turn to medium-low heat for 7 minutes. During the whole process, be careful to prevent the buffer from evaporating and keep the sections moist. Cool naturally and wash the slides three times with PBS (pH 7.4) on a shaker for 5 minutes each time.

[0057] (2) Place the slices in 3% hydrogen peroxide and incubate at room temperature in the dark for 25 minutes to block endogenous peroxidase, then wash with PBS (pH 7.4) on a shaker three times, 5 minutes each time. Use an immunohistochemical pen to mark the location of the tissue, add 3% bovine serum albumin to evenly cover the tissue, and block at room temperature for 30 minutes.

[0058] (3) Discard the blocking solution, add the diluted primary antibody, and incubate the sections in a black humidified box at 4°C overnight. Recover the primary antibody and wash the slides with PBS (pH 7.4) on a shaker three times for 5 minutes each time. Add the secondary antibody of the corresponding species and incubate at room temperature for 50 minutes. Discard the secondary antibody and wash the slides with PBS (pH 7.4) on a shaker three times for 5 minutes each time. Add the freshly prepared DAB color developing solution and immediately observe under a microscope to control the color development time. Positive results will be marked as brown-yellow. Then rinse with running water to stop the color development.

[0059] (4) Restain with hematoxylin for 30 seconds, rinse with running water for 5 minutes, dehydrate, make transparent and seal the slides, and examine under a microscope and take photos.

[0060] XI. Immunofluorescence

[0061] (1) Bake the frozen sections in a 37°C oven for 10-20 min to control the moisture. Fix them in 4% paraformaldehyde for 30 min and wash them three times in PBS on a decolorizing shaker for 5 min each time.

[0062] (2) Place the tissue sections in a repair box filled with EDTA antigen repair buffer in a microwave oven for antigen repair. Heat at medium heat for 8 minutes, stop heating for 8 minutes, and switch to medium-low heat for 7 minutes. During this process, prevent excessive evaporation of the buffer and do not allow the slides to dry out. After cooling naturally, place the slides in PBS and wash them on a decolorizing shaker 3 times, 5 minutes each time.

[0063] (3) After the slices are slightly dried, use a histochemical pen to draw a circle around the tissue (to prevent the antibody from flowing away), dry the PBS, add BSA, and block with sheep serum for 30 minutes.

[0064] (4) Gently shake off the blocking solution, add primary antibody prepared in a certain ratio with PBS onto the slices, and place the slices flat in a humidified chamber and incubate at 4°C overnight.

[0065] (5) Wash the slides three times in PBS on a decolorizing shaker for 5 minutes each time. After the sections are dried slightly, add a fluorescent secondary antibody of the same species as the primary antibody to cover the tissue and incubate at room temperature for 1 hour in the dark.

[0066] (6) Wash the slides in PBS on a decolorizing shaker three times, 5 min each time. After the slides are dried slightly, add DAPI staining solution in the circle and incubate at room temperature for 10 min away from light.

[0067] (7) Wash the slides in PBS on a decolorizing shaker three times, 5 min each time. Add an autofluorescence quencher to the circle for 5 min and rinse with running water for 10 min.

[0068] (8) Add anti-fluorescence quenching sealing agent to seal the slices, and observe the slices under a fluorescence microscope to collect images.

[0069] The present invention is further described below in conjunction with the accompanying drawings and specific examples, but the examples do not limit the present invention in any form. Unless otherwise specified, the reagents, methods and equipment used in the present invention are conventional reagents, methods and equipment in the art.

[0070] Unless otherwise specified, the reagents and materials used in the following examples are commercially available.

[0071] Example 1: Effects of C3G intervention in the parent generation on glucose and lipid metabolism in the offspring

[0072] 1. HFHF-induced glucose metabolism disorder in F1 mice and the improvement effect of C3G

[0073] Using IVF technology, F0 generation mice and wild-type superovulated female mice were used to obtain F1 generation male and female mice with four dietary backgrounds: ND+Vehicle, HFHF+Vehicle, HFHF+C3G, and HFHF+Met, to explore the paternal intergenerational inheritance induced by HFHF diet and the effect of C3G intervention. Figure 2 As far as we know, the weight of F1 mice with F0:HFHF diet background was significantly higher than that of mice with F0:ND diet background before the experiment. In order to investigate the susceptibility of F1 mice to glucose and lipid metabolism disorders caused by high-fat diet, F1 mice were divided into ND diet and HFD diet treatment. The results showed that the aforementioned weight difference gradually became insignificant after 8 weeks of ND feeding; continuing HFD diet would lead to more and more significant weight difference, while the parent C3G diet could not reverse the weight difference, while the weight of the parent Met-treated mice showed effective control ( Figure 2 BC). At the end of the experiment, the weight of the mouse organs was measured and the organ coefficient was calculated. The results are shown in Table 2. In male mice, the HFHF diet of the father generation did not cause changes in the weight of the main organs. The OGTT test showed that in female mice, the administration of HFD to the F1 generation led to a decrease in glucose tolerance in mice with a HFHF background in the F0 generation, while the F0:Met diet had the effect of inhibiting this upward trend ( Figure 2 EF). The ITT test on mice found that male F1 mice showed significant insulin resistance even when fed a normal diet or a high-fat diet due to their parents' HFHF+Vehicle diet, and this insulin resistance was significantly improved under the intervention of C3G and Met from their parents ( Figure 2 GH).

[0074] Table 2: F1 male organ weights

[0075]

[0076]

[0077] In the above table, Vehicle is the solvent, and normal saline is used.

[0078] 2. HFHF-induced lipid metabolism disorders in F1 mice and the improvement effect of C3G

[0079] like Figure 3 As shown in A, the total plasma cholesterol of male mice did not change with the change of the parent's diet, and there was no significant difference between the groups. In female mice, when the offspring were given a high-fat diet, the mice with a HFHF diet background of the parents showed a significant increase in total plasma cholesterol, indicating that the offspring with a HFHF diet background of the parents were more sensitive to the high-fat diet and were prone to high cholesterol levels, and that both C3G and Met treatment of the parents had no effect on alleviating this increase. The results of plasma triglycerides showed that in male mice, the HFHF diet of the parents did not cause abnormal total triglycerides in the offspring; while in female mice, the HFHF diet of the parents caused an increase in total triglycerides in the offspring mice, but the difference was not significant ( Figure 3B). The results of high-density lipoprotein measurement showed that the plasma HDL-C level of male F1 mice was not affected by the dietary background differences of their parents, while the HDL-C of female F1 mice whose parents were fed HFHF+C3G diet background was significantly increased under the influence of high-fat diet ( Figure 3 C). There was no significant difference in low-density lipoprotein in F1 male mice, but in the normal diet group of female F1 mice, the LDL-C levels of mice with HFHF diet background in their parents all decreased ( Figure 3 D).

[0080] The results of histopathological analysis of mouse liver were as follows Figure 3 As shown in EF, even if male F1 mice were given a normal ND diet, mice with HFHF background in the father generation still had a large number of fat vacuoles, and the NAS score increased significantly, indicating that the HFHF diet of the father generation caused the intergenerational inheritance of non-alcoholic fatty liver disease in male offspring; while the offspring of mice whose fathers were treated with C3G and Met had intact hepatocyte morphology, reduced fat vacuoles, and significantly reduced NAS scores. The study also found that when F1 male mice were given a HFD diet, all groups showed signs of non-alcoholic fatty liver disease, and this symptom was not improved by C3G or Met treatment in the father generation. In addition, the pathological outcomes caused by the above intergenerational inheritance were not found in female mice. The above results show that HFHF in the father generation can cause intergenerational inheritance of non-alcoholic fatty liver disease, so that male offspring will also show signs of non-alcoholic fatty liver disease under normal diet conditions, and the treatment of C3G and Met in the father generation significantly inhibited the progression of non-alcoholic fatty liver disease in the offspring.

[0081] Example 2: Improvement of uterine hypertrophy in offspring by C3G intervention in the father generation

[0082] 1. Improvement of uterine morphology and volume of F1 offspring female mice by C3G intervention in F0 generation

[0083] The uterus and ovaries are important components of the female reproductive system and are closely related to reproductive health. As shown in Table 3, in female mice, the HFHF diet of the father caused an increase in the weight of the uterus in the offspring, and the C3G and Met treatment of the father significantly improved this trend. The unilateral morphological images and volume statistics of the uterus of F1 offspring mice showed that the HFHF diet of the father caused uterine hypertrophy in its F1 offspring, and the intervention of C3G and Met of the father significantly improved the phenomenon of uterine hypertrophy in F1 offspring, and the size of its unilateral uterus was similar to that of the F1 offspring of F0:ND+Vehicle ( Figure 4 AB).

[0084] The uterus is composed of an outer serosa, a middle muscularis, and an inner mucosa (endometrium). Figure 4C) Pathological analysis showed that compared with the F1:ND offspring of F0:ND+Vehicle, some offspring of F1:ND on F0 HFHF diet showed increased endometrial thickness, and uterine glands were observed to be enlarged or irregular in shape; C3G intervention in F0 improved the phenomenon of endometrial thickening in F1:ND offspring. Some epithelial cells in the endometrium of F1:HFD offspring lost their normal columnar shape, reflecting atypical hyperplasia, while most endometrial epithelial cells in F1:HFD offspring treated with C3G and Met in F0 maintained neat columnar arrangement. Statistical analysis of the myometrium and endometrium areas of F1 offspring mice showed that the endometrium area of ​​F0:HFHF+Vehicle offspring increased compared with F0:ND+Vehicle F1:ND offspring; compared with F0:HFHF+Vehicle F1:ND offspring, the endometrium area of ​​F0 C3G and Met-treated F1:ND offspring was significantly reduced, and the myometrium area also tended to decrease ( Figure 4 D). For the F1 offspring fed HFD, the endometrial area and myometrial area of ​​the F1:HFD offspring treated with C3G and Met in the F0 generation were significantly reduced ( Figure 4 D).

[0085] Table 3: F1 female organ weights

[0086]

[0087] In the above table, Vehicle is the solvent, and normal saline is used.

[0088] 2. C3G intervention in F0 generation improves HFHF-induced uterine cell proliferation in F1 offspring female mice

[0089] 1. Parental C3G intervention improves PCNA overexpression in the offspring uterus

[0090] Proliferating cell nuclear antigen (PCNA) is a commonly used marker for characterizing cell proliferation. Therefore, immunohistochemistry was used to detect the expression of PCNA in F1:ND offspring to characterize the proliferation level of uterine cells. Compared with the F1:ND offspring of F0:ND+Vehicle, the HFHF diet of the F0 generation significantly increased the expression level of PCNA in the uterus of the F1:ND offspring; compared with the F1:ND offspring of F0:HFHF+Vehicle, the C3G intervention of the F0 generation significantly alleviated the overexpression of PCNA in the uterus of the F1:ND offspring ( Figure 5AB). To further clarify the specific location of the F0 generation where C3G intervention alleviates excessive proliferation of uterine cells, statistical analysis was performed on the PCNA-positive expression areas of the endometrial stroma, endometrial epithelium, and uterine myometrium. Figure 5 B The results showed that the HFHF diet of the F0 generation significantly caused an increase in the PCNA-positive expression area of ​​the myometrium of the F1:ND offspring female mice. At the same time, C3G and Met intervention of the F0 generation significantly improved the PCNA-positive expression area of ​​the myometrium of its F1:ND offspring.

[0091] 2. C3G intervention in the parental generation improves the overproliferation of myometrial cells in the offspring

[0092] Through quantitative analysis of the PCNA positive expression area in different regions, it was found that the myometrium may be the site where the HFHF diet causes excessive proliferation of uterine cells in offspring. To further confirm the proliferation site of PCNA in the uterus of mouse offspring, α-smooth muscle actin (α-SMA) was used to locate uterine smooth muscle cells to characterize the mature myometrium. The results showed that the F0 generation HFHF diet led to a significant increase in the PCNA fluorescence intensity in the myometrium of F1 offspring mice, while C3G and Met intervention in the F0 generation significantly improved the fluorescence intensity of PCNA in the myometrium ( Figure 6 AB). Therefore, the HFHF diet of the F0 generation leads to excessive proliferation of myometrial cells in the F1:ND offspring female mice, and C3G and Met intervention of the F0 generation can restore the normal proliferation level of myometrial cells in the F1:ND offspring.

[0093] In summary, the present invention first discovered that a high-fat, high-fructose diet of the parent generation can lead to insulin resistance and non-alcoholic fatty liver disease in the F1 offspring, and cause uterine hypertrophy in the F1 offspring, which has adverse intergenerational genetic effects on the F1 offspring. C3G intake by the parent generation can significantly improve insulin resistance in the F1 offspring, alleviate the phenomenon of non-alcoholic fatty liver disease, and alleviate uterine hypertrophy in the F1 offspring. The intake of anthocyanins by the parent generation can protect the health of the offspring's sugar and lipid metabolism and reproductive health, and can effectively improve adverse intergenerational genetic effects, providing new ideas for functional foods that prevent adverse intergenerational inheritance.

[0094] The above embodiments are preferred implementation modes of the present invention, but the implementation modes of the present invention are not limited to the above embodiments. Any other changes, modifications, substitutions, combinations, and simplifications that do not deviate from the spirit and principles of the present invention should be equivalent replacement methods and are included in the protection scope of the present invention.

Claims

1. Application of cyanidin-3-O-glucoside in the preparation of functional foods for preventing adverse intergenerational inheritance.

2. The use according to claim 1, characterized in that: The adverse intergenerational inheritance is glucose and lipid metabolism disorder in the offspring or uterine hypertrophy in the female offspring.

3. The use according to claim 2, characterized in that: The offspring glucose and lipid metabolism disorder is selected from one of the following: glucose intolerance; insulin resistance; abnormal total cholesterol, total triglycerides, high-density lipoprotein cholesterol and low-density lipoprotein cholesterol in plasma; and non-alcoholic fatty liver.

4. The use according to claim 2, characterized in that: The female offspring uterine hypertrophy is selected from one of the following: an increase in the volume of the offspring uterus; an increase in the area of ​​the uterine myometrium; an increase in the area of ​​the endometrium; and excessive proliferation of offspring uterine cells.

5. A functional food for preventing adverse intergenerational inheritance, characterized in that: The functional food contains cyanidin-3-O-glucoside.

6. The functional food according to claim 5, characterized in that: Also included are acceptable excipients.

7. The functional food according to claim 5, characterized in that: The functional food is prepared into a type selected from the following: tablets, powder, pills, capsules, granules, dripping pills, and oral liquid.

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