Therapeutic Effect And Mechanism Of Ligusticum Wallichii Extraction And The Prescription On Testosterone- Induced Hair Loss in C57BL/6 Mice Ⅱ
Oct 31, 2022
3 Results and discussion
3.1 Hair growth in mice
Hair growth in mice consists of telogen, catagen and anagen phases, known as the anagen cycle, and the depilated skin color deepens as hair follicles transition from telogen to anagen. The hair follicles of C57BL/6 mice contain melanocytes, which can synthesize a large amount of melanin during the growth phase, and the skin is black; the synthesis of melanin decreases in the regression phase, and the skin is gray; The hair follicles of 6-8 week old C57BL/6 mice enter the telogen phase, therefore, the model of androgenetic alopecia can be easily established by administering testosterone during this period.
It can be seen from Figure 1 that Day 1 is the back skin of the mice immediately after the end of modeling 1 day, the color is gray and dull, and the hair growth is in a state of rest. After stimulation with different traditional Chinese medicine extracts, it can make hair grow into the growth phase. On the 4th day of administration, the back of the mice in each administration group was
The hair on the top did not grow significantly, but the skin gradually darkened. After 8 days of administration, the hair growth tended to be obvious, and only a few areas of the mice in the model group grew hair, while the mice in the administration group had different degrees of hair growth, and the promotion effect of the low-dose chuanxiong group was compared with other groups. The drug group was more obvious, indicating that minoxidil, high and low doses of chuanxiong and compound can significantly shorten the transition time of hair follicles from telogen to anagen and the time from entering anagen to hair growing out of the epidermis. After 12 days, only a small amount of hair grew in the mice in the model group, most of the back area was still bare, the skin was white, and the hair growth was in the telogen phase. However, the hair on the back of the mice in each administration group basically covered the entire depilation area on the back, and the color was gray-black and shiny. Some mice developed gray hair on their backs, and some mice treated with high-dose Chuanxiong extract showed hair loss on the upper neck and lower buttocks.
3.2 Determination of hair length and hair coverage
It can be seen from Table 1 that on the 8th and 10th days of treatment, the mice in the model group had no back hair or the length could not be measured, while the treatment group grew shorter hairs (3 and 5 mm). On the 12th day, different degrees of new hair grew on the backs of mice in all groups, and the ratio of the length of new hair in each administration group to the length of hair in the model group (EE1) was used to evaluate the effect of each formula on promoting hair recovery. The order of promotion effect: low-dose Chuanxiong group>minoxidil group>compound formula group> Chuanxiong high-dose group. Among them, the EE1 of the Chuanxiong low-dose group reached 3.4, followed by the minoxidil and Angelica low-dose groups, the EE1 reached 3.2. The results showed that the Chuanxiong extract and the compound formulation had obvious effect on promoting hair growth, and the low-dose Chuanxiong group had better effects than the higher-dose group, suggesting that the effect of Chinese herbal extracts on promoting hair growth was related to the concentration.
It can be seen from Table 2 that on the 4th day of treatment, no hair grew in the alopecia area of the mice in all groups. On the 8th day, a small part of the back of the mice in the model group was covered with hair, while more than half of the back of the mice in the administration group was covered with hair. On the 12th day, all mice covered their backs to varying degrees.
Cover the hair, calculate the percentage of hair coverage (%) based on the ratio of the newly-born hair coverage area of the mice in each group to the area of the total shaved area, and use the ratio of the hair coverage percentage of the mice in each administration group and the hair coverage percentage of the model group on the 12th day. The ratio (EE2) was used to evaluate the promoting effect of each formula on hair recovery. The order of promotion effect: low-dose Chuanxiong group> compound formula group> minoxidil group> Chuanxiong high-dose group. Among them, the EE2 of the Chuanxiong low-dose group reached 5.4, followed by the compound formula group, which reached 5.3.
Cistanche for improving testosterone
3.3 Determination of skin moisture, oil and temperature
Decreased skin surface moisture, increased oil, and elevated temperature are major features of AGA [3].
. It can be seen from Figure 2 that compared with the blank group, the moisture value of the back skin of the mice in the model group decreased (P < 0.05), the oil content increased (P < 0.05), and the temperature also increased. The model was built successfully. After being treated with different formulas, the water, oil and temperature of the mice in the administration group recovered to different degrees. For moisture, the skin moisture value of mice in each administration group returned to the level of the blank group, and the high-dose Ligusticum chuanxiong group had the best effect. The combination group was more obvious than the minoxidil group; the skin temperature in each administration group returned to normal. The results showed that both the Chuanxiong extract and the compound have the effects of reversing the decrease in skin moisture content, excessive oil secretion and restoring the normal temperature of the skin, thereby preventing hair loss and promoting hair growth. Dil works better.
3.4 Observation of pathological sections
The results of histopathological examination (Fig. 3) showed that the number of hair follicles in the skin of the mice in the blank group was more, and some of them were in the growth phase. Compared with the blank group, the longitudinal sections of the skin of the mice in the model group showed shrinkage and atrophy of hair follicles, accompanied by a decrease in the total thickness of the skin, and the morphology of the hair follicles showed that they were still in the spontaneous resting phase, suggesting that the AGA model was successfully developed in C57BL/6 mice. Establish. The AGA model is characterized by the gradual miniaturization of hair follicles. The miniaturized hair follicles stay in the telogen phase and cannot enter the growth phase of the next cycle. It is also related to the gradual reduction of hair diameter, which will cause the terminal hair to become vellus hair [6]. Compared with the model group, the skin thickness of the mice in each administration group recovered to the level of the blank group. After intervention with different Chinese herbal extracts, the number and diameter of hair follicles in mice were significantly increased. Among them, the mice in the high-dose Chuanxiong group had the largest number of skin hair follicles hyperplasia and the volume of the hair follicles was relatively large, which was comparable to the minoxidil group, which helped to grow more hair and form thicker hair shafts. In addition to the hair follicle morphology, compared with the model group, the significant increase in melanin in each administration group could also indicate that the hair follicle had entered an active growth phase.
3.5 Detection of VEGF, α-MSN and TGF-β1 contents in mouse skin In this experiment, the expression of VEGF, α-MSN and TGF-β1 was used to evaluate the therapeutic effect of traditional Chinese medicine extract on hair loss, and to try to explain Chuanxiong from the molecular level. The mechanism of anti-dropping effect of traditional Chinese medicine. Hair renewal involves changes in hair follicle morphology and its microvascularization, and alopecia is associated with reduced hair follicle microvascularization. VEGF is one of many angiogenic factors, has a strong pro-angiogenic activity, can promote the formation of microvessels in the process of hair follicle differentiation, so it is of great significance for hair follicle hair renewal [14]. For the expression of VEGF (vascular endothelial growth factor) (Fig. 4), after treatment with different drugs, the level of VEGF in the skin of mice was significantly increased compared with the model group, among which the low-dose group of Chuanxiong had the most obvious effect, followed by the high-dose of Chuanxiong. group and minoxidil group, while the compound formula group had less obvious effect. α-MSH and TGF-β1 are secreted by anagen hair bulbs, which are important for the maintenance of hair follicle immune forgiveness [15]. The immune amnesty of hair follicles is a dynamic, periodic, and relative amnesty state, which only exists in the growth phase of the hair follicle. The permanent part enters the resting period [16].
Therefore, the restoration and maintenance of immune forgiveness determines the onset and duration of the growth phase of the hair follicle and is Essential for maintaining hair growth. As shown in Figure 5, compared with the model group, the content of α-MSN in the skin of mice in different administration groups was increased, and the content levels were in descending order: Chuanxiong low-dose group > compound formula group > > positive group > Chuanxiong high-dose group. For the expression of TGF-β1 (Fig. 6), the order of content level from high to low was: compound formula group> Chuanxiong low-dose group> Minoxidil group> Chuanxiong high-dose group. Overall, the hair growth-promoting effect of the Chuanxiong extract and the complex formulation may be related to its enhanced restoration and maintenance of hair follicle immune amnesty and angiogenesis in alopecia areas.
