Reducing blood lipid behavior in vivo based dark brick tea intake

ABSTRACT

Dark brick tea, a unique kind of dark tea has a history of more than 100 years in China. In this paper, 1 and 10 years old dark brick tea were used to regulate blood lipid. The results showed that dark brick tea could significantly reduce the total cholesterol (TC) and triglyceride (TG) levels in serum and liver homogenate. It also significantly increased the level of high-density lipoprotein cholesterol (HDL-C). In addition, dark brick tea can enhance the activities of superoxide dismutase (SOD) and glutathione peroxidase (GSH-Px) in rats, reduce the content of malondialdehyde (MDA), promote the activities of lipoprotein lipase (LPL) and liver lipase (HL) in rats, and effectively reduce blood lipid levels. In conclusion, the experimental results of this study will provide a scientific basis for the lipid-lowering effects of black brick tea with a theoretical basis for its further development and utilization in the form different products.

KEYWORDS:

• Dark brick tea
• green tea
• blood lipid
• mechanism
• utilization

1. Introduction

As verified by numerous research reports in China and abroad, hyperlipidemia and lipid metabolism disorder can induce atherosclerosis which is an important causative factor of coronary heart diseases (Lv et al., 2021; Wang et al., 2022). If the blood lipid level will be kept under control through dietary treatment or drugs, the progress of atherosclerosis can be postponed or prevented and will thus aid in reducing the morbidity and mortality due to coronary heart diseases (He et al., 2022; Song et al., 2021). However, current hypolipidemic drugs may trigger various side effects, making them unsuitable for long-term use, or they may fail to attain a satisfactory curative effect due to their weak effects (Hua et al., 2021). Therefore, the present-day research on prevention and treatment of cardiovascular diseases should seek for natural, effective, and safe hypolipidemic drugs or foods.

Tea polyphenols and pigments have been found to resist lipid peroxidation, prevent hyperlipidemia, and reduce the incidence of cardiovascular and cerebrovascular diseases (Bing et al., 2022; Wang et al., 2022). Polyphenols can regulate lipid metabolism and hence play an important role in lowering blood lipid (Pan et al., 2022). Tea polyphenols can reduce the total cholesterol (TC) and triglyceride (TG) levels in blood serum and increase the high-density lipoprotein cholesterol (HDL-C) level, more efficiently than that of inositol nicotinate (Dong et al., 2022; Duarte et al., 2021; Li et al., 2021). This implies that tea polyphenols can be used to prevent and treat hyperlipidemia. The experiment on tea pigment capsules regulating blood lipid metabolism revealed that it is a safe and effective drug for regulation of blood lipid metabolism, and can exert a hypolipidemic effect. However, that also came up with some side effects as reported previously (Yao et al., 2021). Hyperlipidemic patients took tea pigment orally by adopting self-control method, and indexes including blood lipid, blood sugar, platelet, liver function, kidney function, blood potassium, sodium, and chlorine, were measured. After administration of tea pigment for months, the levels of TC, TG, and HDL-C in blood serum were all reduced (Maron et al., 2003; Wu et al., 2021; Xu et al., 2022). Tea pigment is an ideal hypolipidemic drug without any toxic side effect after long-term administration (Deng et al., 2021; Zhu et al., 2021). According to evaluatiing the results of Pu’er tea’s function in regulating blood lipid, sun-dried green tea and Pu’er tea were found to effectively inhibit the increase in blood lipid in mice fed with high-fat diets maintaining all blood lipid indexes (TC, TG, and LDL) in the normal ranges (Yue et al., 2022). The effect of Pu’er tea was slightly better than that of sun-dried green tea, and its effect on TG was stronger than that on TC. Pu’er black tea showed a poorer hypolipidemic effect than Pu’er green tea, as revealed by the paired observation of their hypolipidemic function for old people. Moreover, Pu’er tea and its polyphenolic oxidative products effectively regulated serum lipid level in high-fat diet fed mice, mainly reflected by the decrease in serum TC and TG levels (Hou et al., 2022; Hu et al., 2022; Ye et al., 2021).

The hypolipidemic function of dark brick tea has been scarcely investigated. With dark green tea as the control material, 1 and 10 years old dark brick tea were used as experimental materials to analyze their chemical components, and further explore their role in regulating blood lipid. This study provides a new direction for green and effective auxiliary hypolipidemic products via further development of the old dark brick tea market.

2. Materials and methods

2.1 Materials

Dark brick tea was provided by Zhaoliqiao Tea Factory, Chibi city, Hubei Province. The “Chuan” brand dark brick tea was divided into the current-year green brick (called 1-year old brick) and 10-year brick (also called 10-year brick) tea. Green tea was used Enshi Grade 4 fried green tea. The positive drug was Xuezhikang produced by Beijing Beida Weixin Biotechnology Co., Ltd.

2.2 Determination of main components

Moisture, water extract, tea polyphenols, amino acids, soluble sugar, and protein were determined in accordance with GB/T 8314–2002 protocol. Theaflavin, thearubigins, and theabrownin were determined following the same procedures as mentioned in the previously published papers (Jia et al., 2022; Li et al., 2022).

2.3 Design of experimental animals

A total of 80 amputated male Wistar rats (SPF grade, weight: 150 ± 5 g) were provided by Hubei Experimental Animal Research Center. The reason for selecting male rats is they are stronger than female, and not easily influenced by environmental factors. Dose selection: Ordinary animal feed was provided by Hubei Experimental Animal Research Center. The formula of high-fat diet was as follows (%): basic feed (78.8), pig fat (10.0), egg yolk powder (10.0), cholesterol (1.0), and bile salt (0.2). Eight groups (with 10 rats in each group) were set up in the experiment, namely, blank control group (NC), model control group (MC), positive control group (XZK, 0.12 g/Kg·bw), 1-year brick tea 5-fold dose group (DBTL, 0.75 g/Kg·bw), 10-fold dose group (DBTM, 1.5 g/Kg·bw), 20-fold dose group (DBTH, 3.0 g/Kg·bw) calculated based on the recommended dose for human body, 10-year brick tea 10-fold dose group (10DBT, 1.5 g/Kg·bw), and green tea 10-fold dose group (GT, 1.5 g/Kg·bw). Seven groups received a high-fat diet except that ordinary feed was given to the NC group. Rats were administered intragastrically at 1 mL/100 g·bw.

2.4 Serum and liver tissue blood lipid indexes measurement

In the experimental environment, rats were fed with the adaptive basic diets for 5 days, and their tail blood was obtained to measure serum TC. Based on the TC level and body weight, the rats were randomly divided into eight groups: NC, MC, XZK, and DBTL, DBTM, DBTM, 10DBT, and GT as explained before. Ten rats were included in each group. The rats were weighed and numbered by the labeling method. The NC was fed with distilled water and ordinary feed. The MC was given distilled water and high-fat diet. The other six dose groups were fed with high-fat diets by intragastric gavage with corresponding test samples. The rats were weighed regularly (twice a week). They were fasted for 12 h at the end of the experiment and then weighed. Blood samples collected from the carotid artery were centrifuged at 2000 rpm and 4°C for 10 min. The serum was separated, and the levels of TC, TG, HDL-C, superoxide dismutase (SOD), malondialdehyde (MDA), glutathione peroxidase (GSH-PX), lipoprotein lipase (LPL), and hepatic lipase (HL) were determined. The fat deposition and organ changes were observed with naked eyes. A total of 12.5% liver homogenate was prepared, and its TC, TG, HDL-C, SOD, MDA, and GSH-PX levels were measured. Serum and liver homogenate levels of TC, TG, HDL-C, SOD, MDA, GSH-PX, total lipase (LPL and HL), and serum lipase were determined through the kit method (Feng et al., 2022; Zhao et al., 2021). The protein in liver tissue homogenate was determined using the Coomassie brilliant blue protein kit (Guo et al., 2021).

2.5 Pathological observation

Pathological observations of liver in MC: After the rats were dissected, 1.0 g of the liver from each group was fixed in 10% neutral formalin solution, which was shaken from time to time to fix the liver completely and fully. The paraffin sections were embedded, followed by hematoxylin and eosin staining. Then, tissue changes were observed under an optical microscope, and the results were recorded by an Olympus universal microscope. Observation of fat cells: About 1 g fat around testicles was obtained and fixed in 10% neutral formalin, which was shaken from time to time to fix the fat completely and fully. The paraffin sections were embedded, HE staining was performed and tissue changes were observed under the optical microscope, and the results were recorded by an Olympus universal microscope.

2.6 Statistical analysis

Each experiment was carried out at least in triplicate, and the results were shown as means ± SD. SPSS 25.0 (SPSS Inc., Chicago, IL, U.S.A.) was used to determine the statistical differences (p < .05) through analysis of variance (ANOVA) and Duncan’s test.

3 Results and discussion

3.1 Main chemical components of dark brick tea

Dark brick tea belongs to post-fermented black tea, which is made of mature new shoots as raw materials. First, new shoots are processed into sun-dried green tea and then processed through pile fermentation, drying, blending, autoclaving, low-temperature long baking, aging, and other processing procedures to impart unique quality characteristics to the dark brick tea. Pile fermentation is the key to the quality formation of dark brick tea. During this process, the main biochemical components of dark brick tea, such as tea polyphenols, protein, and amino acids, change dramatically. However, the contents of soluble sugar and tea saponin show no considerable change, but those of theaflavin, thearubigins, and theabrownin increase significantly. As shown in Table 1, the contents of tea polyphenols, protein, and amino acids in 1-year brick tea were 8.01%, 1.52%, and 1.52% respectively, which were lower than those in ordinary fried green tea, whereas high contents of theaflavin (0.08%), thearubigins (2.27%), and theabrownin (3.48%) were observed. Compared with those in 1-year dark brick tea, the contents of tea polyphenols, theaflavins, and thearubigins in the 10-year brick tea, which went through multi-year storage and aging, continually decreased by 3.81%, 0.03%, and 3.85% respectively, the content increased by 3.85%, whereas the contents of amino acids (1.31%) and proteins (1.35%) changed slightly. Compared with that in green tea (41.76%), the content of water extracts in 1- and 10-year brick tea declined significantly by 20.82% and 17.03%, respectively. The content of water extracts in 10-year brick tea changed minimally compared with that in 1-year brick tea. This finding means that through storage, aging, and fermentation, several of its components, such as tea polyphenols and their oxidation products, namely, sugars and proteins, changed and were possibly converted into other effective substances. This may be because the extracellular enzymes produced by Aspergillus Niger in the process of pile fermentation can catalyze the decomposition of insoluble organic matter in tea into soluble matter, and catalyze the hydrolysis of biological macromolecular organic matter into small molecular organic matter that is easy for microbial absorption and utilization (Yang et al., 2021).

Table 1. Main biochemistry components in green tea and dark brick tea (%).

	Moisture	water extracts	tea polyphenols	amino acids	Soluble sugar	protein	Tea saponin	Theaflavin	Thearubigins	Theabrownin
1-year brick tea	10.58	20.82	8.01	1.52	4.99	1.95	4.65	0.08	2.27	3.48
10-year- brick tea	10.21	17.03	3.81	1.31	4.39	1.35	4.69	0.03	0.72	3.85
Green tea	6.05	41.76	26.25	4.98	5.49	2.67	3.93	—	—	—

3.2 Effect of dark brick tea on the auxiliary hypolipidemic effect of experimental rats

3.2.1 Effect of dark brick tea on the body weight of experimental rats

The experimental results (Table 2), showed that at the beginning of the experiment, no significant difference was observed in the initial average weight among different groups of animals. But the difference in the final average weight was significant, the weight difference between MC and other groups was significant (P < .01). Compared with the MC, the weight loss of rats in the different dose groups of dark brick tea was evident, and the weight of rats declined with the increase in dose. Compared with the body weight of GT, the difference between 10DBT and DBTH was significant (P < .01), which indicates the better weight-reducing effect of dark brick tea than green tea. In this experiment, the different dose groups of dark brick tea exhibited a dose-dependent weight-reducing effect, where the 10-year brick tea exhibited the best effect.

Table 2. Effect of dark brick tea on body weights.

Group	n	Initial body weight (0d)	Final body weight (42f)	Significance
				P < .05	P < .01
NC	10	167.68 ± 6.92	290.16 ± 29.99	a	A
MC	10	167.10 ± 5.52	402.54 ± 30.18	f	D
XZK	10	167.80 ± 11.00	364.46 ± 23.76	e	C
DBTL	10	167.99 ± 5.59	345.66 ± 29.40	cde	BC
DBTM	10	167.41 ± 6.72	335.70 ± 16.97	bcd	BC
DBTH	10	166.24 ± 5.93	321.61 ± 31.75	bc	AB
10DBT	10	164.62 ± 5.35	318.51 ± 32.97	b	AB
GT	10	166.24 ± 6.56	360.36 ± 22.14	de	C

3.2.2 Auxiliary hypolipidemic effect of dark brick tea on experimental rats

The results (Figure 1a), demonstrated a significant difference in TC levels between the serum and liver homogenate of MC compared to other groups (P < .01). The TC levels in the 10DBT group showed no significant difference from those in NC. In comparison to XZK, the serum TC levels in DBTL and DBTM exhibited nonsignificant differences, while DBTH and 10DBT significantly decreased with an extremely notable level of distinction (P < .01). Therefore, DBTH and 10DBT displayed superior efficacy over XZK in reducing serum TC levels. Furthermore, when compared to GT, dark brick tea (DBTM, DBTH, and 10DBT) significantly differed in their ability to lower TC levels (P < .01), indicating a more pronounced effect than green tea. Amongst different dose groups of dark brick tea, there was a clear reduction observed in both serum TC and liver homogenate TC levels among experimental rats. This effect was found to be dose-dependent with the highest effectiveness achieved by the 10-year brick tea sample.

Figure 1. Effect of dark brick tea on serum and liver TC (a), TG (b) and HDL-C (c) level.

3.2.3 Effect of dark brick tea on serum TG and liver tissue homogenate TG levels in experimental rats

The results (Figure 1b), revealed significant differences in serum TG and liver homogenate TG levels between the MC group and the other groups (P < .01). The 10DBT group exhibited no significant difference in serum and liver homogenate TG levels compared to the NC group, indicating a desirable effect of reducing TG levels with 10-year-old brick tea. SNK and DBTL showed no significant difference in serum TG level, while DBTM, DBTH, and 10DBT displayed a substantial decline that was statistically significant (P < .01). Thus, dark brick tea had a better TG-lowering effect than green tea. Comparisons showed that the different dose groups of dark brick tea can remarkably reduce the serum TG and liver homogenate TG levels in experimental rats, with the effect being dose-dependent, where the 10-year brick tea demonstrated the most pronounced impact.

3.2.4 Effect of dark brick tea on serum HDL-C and liver tissue homogenate HDL-C levels in experimental rats

As revealed by the experimental results (Figure 1c), the serum HDL-C level in MC group significantly differed from those in NC, DBTM, DBTH, and 10DBT (P < .01). Similarly, the liver homogenate HDL-C level in the MC group significantly differed from that in the SNK, GT, and different dose groups of dark brick tea (P < .01). While no significant difference was observed in serum HDL-C levels between DBTM and DBTH compared to NC, it was significantly different from 10DBT (P < .01). Furthermore, it is worth noting that an evident effect on elevating serum HDL-C levels was observed. The liver homogenate HDL-C level in NC was significantly different from those in different dose groups of dark brick tea(P < .01). Furthermore, when compared to XZK, the serum HDL-C levels significantly decreased in the DBTM, DBTH, and 10DBT groups (P < .01). Similarly, a significant disparity was observed between GT and DBTM, DBTH, and 10DBT regarding their respective HDL-C levels (P < .01). These comparisons demonstrate that different doses of dark brick tea can effectively elevate both serum HDL-C and hepatic homogenate HDL-C levels in experimental rats. Moreover, this effect is dose-dependent with 10-year brick tea exhibiting the most pronounced impact. This experiment was an animal experiment on auxiliary hypolipidemic functions, which was carried out completely in accordance with Standard for Functional Evaluation Procedures and Testing Methods of Health Food. The results hinted that different doses of dark brick tea can notably reduce serum TC and TG levels and liver homogenate TC and TG levels in experimental rats and evidently increase their serum HDL-C and liver homogenate HDL-C levels in the rats. In the NC group, the deposition of fat was observed predominantly in vivo, with the liver exhibiting a yellow and white appearance. Additionally, the upper oil layer of liver homogenate appeared thick. The alterations in indices for DBTH and 10DBT were significantly superior to those for XZK and GT, suggesting that dark brick tea possesses an auxiliary hypolipidemic function. Notably, 10-year brick tea demonstrated the most effective auxiliary hypolipidemic effect among all tested samples.

3.3.1 Effect of dark brick tea on serum and liver tissue homogenate SOD levels in experimental rats

The experimental results (Figure 2a), showed that compared with that in MC, the serum SOD levels in other groups were significantly different (P < .01). In comparison with that in NC, the serum SOD levels in SNK, DBTL, DBTM, and DBTH were not significantly different, but a significant difference was observed from that in 10DBT. The serum SOD level in NC was highly significantly different from those in DBTH and 10DBT (P < .01), and the effect of elevating the serum SOD level was evident. The serum SOD level in XZK was significantly different from those in DBTH and 10DBT (P < .01). Thus, the effect of increasing the SOD level in DBTH and 10DBT was better than that in SNK. Compared with GT, a significant difference was observed between DBTH and 10DBT (P < .01). Evaluating the effects of different dose groups of dark brick tea, extremely significant differences were observed from the SOD levels in DBTH and 10DBT (P < .01). Similarly, the serum SOD levels in DBTL and DBTM were found to be significantly differed from those in DBTH and 10DBT (P < .01). The effect was dose dependent, and 10-year dark brick tea attained the best effect.

Figure 2. Effect of dark brick tea on serum and liver SOD (a), MDA (b) and GSH-PX (c) level.

3.3.2 Effect of dark brick tea on serum and liver tissue homogenate MDA levels in experimental rats

According to experimental results (Figure 2b), the levels of serum MDA and liver homogenate MDA in the MC group were found to be significantly different from those observed in other groups (p < .01). These results indicate that each dose group exhibited a significant reduction in MDA activity within the experimental rats. Compared with the serum MDA level in NC group, no significant difference was observed from those in XZK, DBTL, DBTM, and GT, but the difference was significant from that in DBTH (P < .05) and highly significant from that in 10DBT (P < .01). Thus, each dose group exerted a good effect on reducing MDA activity in experimental rats, especially in DBTH and 10DBT. Compared with that in XZK, the serum MDA levels in DBTM and DBTH were significant (P < .05), and the difference from those in DBTH and 10DBT groups was extremely significant (P < .01). Thus, DBTM, DBTH, and 10DBT were superior to XZK in reducing the MDA level. The liver homogenate MDA level in GT was highly significantly different from that in 10DBT (P < .01). Similarly, a significant difference was observed in the liver homogenate MDA level except DBTL (P < .05), but found not to be significantly different among other groups. Therefore, the serum MDA and liver homogenate MDA levels in experimental rats were evidently reduced, with the effects to be dose dependent. The10-year-old brick tea achieved the best effect.

3.3.3 Effect of dark brick tea on serum and liver tissue homogenate GSH-PX levels in experimental rats

The experimental results (Figure 2c), showed highly significant differences among the serum GSH-PX levels in MC and those in NC, DBTM, DBTH, and 10DBT (P < .01) groups. Therefore, DBTM, DBTH, and 10DBT can remarkably improve serum GSH-PX activity in experimental rats. Compared with that in NC, the serum GSH-PX levels in DBTM, DBTH, and 10DBT were not significantly different. As for the liver homogenate GSH-PX level, NC did not significantly differ from DBTH and 10DBT. The GSH-PX level in XZK was significantly different from those in DBTH and 10DBT (P < .01). Also, compared with that in GT, the GSH-PX levels in DBTH and 10DBT were significantly different (P < .05). Furthermore, comparisons of different dose groups of dark brick tea showed no significant difference in the other groups except in the liver homogenate GSH-PX in DBTL (P < .05). Hence, serum and liver homogenate GSH-PX levels in experimental rats were evidently elevated. The effect was dose dependent, and 10-year brick tea achieved the best effect.

3.3.4 Effect of dark brick tea on serum LPL and HL activities in experimental rats

Effect on serum LPL activity in experimental rats: The experimental results (Figure 3), showed that all groups were extremely significantly different from MC in the serum LPL level (P < .01), which indicates that all dose groups can significantly improve the serum LPL activity in experimental rats. No significant differences were observed between NC and 10DBT in terms of the serum LPL level. Compared with XZK, the LPL levels in DBTM, DBTH, and 10DBT were significantly different (P < .01). Thus, medium- and high-dose groups of dark brick tea exerted better effects on elevating the LPL activity than SNK. DBTM, DBTH, and 10DBT were extremely significantly different from GT (P < .01), which implied that the LPL level was elevated better by dark brick tea than by green tea. Through comparisons of different dose groups of dark brick tea, DBTL was extremely significantly different from DBTM, DBTH, and 10DBT (P < .01), and it can evidently increase the serum LPL level in experimental rats. The effect was dose dependent, and 10-year dark brick tea exerted the best effect.

Figure 3. Effect of dark brick tea on LPL, HL level of serum and liver.

Effect on HL activity in experimental rats: The experimental results given in the (Figure 3), exhibited that the serum HL level in MC showed extremely significant differences from those in NC and different dose groups of dark brick tea (P < .01). Therefore, the serum HL activity in experimental rats can be significantly improved by different doses of dark brick tea. The HL level in XZK was highly significantly different from those in DBTM, DBTH, and 10DBT (P < .01), which manifested that the medium- and high-dose groups of dark brick tea exerted a better effect on elevating the HL level than SNK. Compared with that in GT, the HL levels in DBTM, DBTH, and 10DBT were significantly different (P < .01), which proved the better effect of dark brick tea on increasing the HL level than green tea. Compared with DBTL and DBTM, the different doses of dark brick tea also displayed significant differences in the HL level (P < .05), which can remarkably increase the serum HL level in experimental rats. The effect was dose dependent, and the 10-year brick tea reached the best effect.

3.3.5 Effect of dark brick tea on liver morphologies of experimental rats

The liver of rats in NC presented a crimson and elastic appearance. In MC, most of the livers turned yellow, and a large amount of fat was precipitated out of the liver homogenate. The livers of rats in other groups were slightly lighter in color. The slice map (Figure 4), showed that in NC, the hepatic lobule was of a distinct structure, with a clear central vein accompanied by radially arranged hepatic cords, normal hepatic sinuses, consistently sized hepatic cells, clear cell nucleus, and uniform cytoplasm. In MC, the normal structure of hepatocytes disappeared, cell boundaries were unclear, hepatic cords were arranged disorderedly, hepatocytes were swollen and denatured in a large area, and the cytoplasm was vacuolar and lightly stained. In other experimental groups, the symptoms were relieved, cell boundaries were evident, and vacuole areas were small. 10DBT showed better liver protection, without any significant difference from NC, which indicated that dark brick tea exerted a good liver protection effect on experimental rats, with the 10-year-old dark brick tea achieving the best effect. The investigation results regarding the auxiliary hypolipidemic mechanism of dark brick tea for experimental rats showed that dark brick tea significantly increased SOD and GSH-Px in serum and liver homogenate of experimental rats, significantly decreased MDA, and enhanced the activity of serum LPL and HL in experimental rats. Liver tissue slices displayed that the normal structure of liver cells in MC of experimental rats disappeared, and the damage degree to the normal structure in each experimental group was lighter than that in MC and mainly manifested in clear cell boundaries and less vacuole area.

Figure 4. Liver cells pathological observations of each group rats.

4. Conclusion

Dark brick tea has been shown to significantly decrease the levels of to TC and TG in both serum and liver homogenate of experimental rats, while also noticeably increasing the level of HDL-C. Therefore, dark brick tea exhibits a dose-dependent auxiliary hypolipidemic effect on experimental rats. Notably, 10-year brick tea demonstrates a superior auxiliary hypolipidemic effect compared to 1-year dark brick tea. Additionally, dark brick tea enhances the activities of SOD and GSH-Px, reduces MDA content, and promotes LPL and HL activities in rats to effectively lower blood lipid levels. This study provides a good data base for the hypolipidemic effect of black brick tea, and provides a scientific basis for the development and utilization of its industry based on the concept of homology of medicine and food. This study will continue to further isolate and identify the components of dark brick tea, and evaluate the lipid-lowering and hypoglycemic effects of dark brick tea through in vitro tests, in order to comprehensively elaborate the pharmacodynamic material basis of green brick tea for lipid metabolic diseases.

Ethics approval

There is no ethics interest related in research. The research was granted by Ethics Committee of Xinyang Normal University.

Disclosure statement

There is no professional or other personal interest of any service and/or company that could be construed as influencing the position presented in the manuscript. The work has not been published before.

Additional information

Funding

This work was financially supported by the Natural Science Foundation of Henan Province (Grant No. 212102110419). The work is kind supported by College of Horticulture and Forestry in Huazhong Agricultural University.