| Objectives Ischemic cerebrovascular disease has the characteristics of high morbidity high mortality and morbidity, and the high rate of relapse. At present, The curative effect of progesterone on cerebral ischemia protection basic research is quite mature at domestic and abroad, and nerve protective effect on cerebral ischemia or traumatic brain injurie has been identified by progesterone. The most representative one is progesterone, and its the side effects of hormone hindered the clinical application. Ischemic brain injury belongs to the scope of TCM "stroke". TCM treatment related diseases has clinical curative effect. A large number of studies have shown that traditional Chinese medicine monomer baicalin, one of the main efficacy components in traditional Chinese medicine radix scutellariae, can not only increase progesterone level but also have a certain role for stroke.This topic research is based on progesterone protection against ischemic brain injury research, combined with related theoretical guidance of Traditional Chinese Medicine. TCM monomer baicalin regulating biological progesterone levels in biological body was regarded as the breakthrough point.The possible mechanism was studied in vitro whether brain protection of baicalin was via influencing progesterone in female MCAO rats.Detailed in the following three parts:l.To clarify the protective effect of baicalin on ischemic brain injury;2.To certify the regulating effect of baicalin on serum progesterone and correlated hormones level in normal and ischemic brain injury rats.3.To explore the possible mechanism of the protective effect of baicalin on ischemic brain injury.MethodsWith vaginal smear method, the adult estrus female SD rats were selected for experiments. The adult estrus female SD rats were selected and divided into normal group, baicalin normal group, model group. The left side of the middle cerebral artery of rats was blocked to establish the permanent middle cerebral artery occlusion(pMCAO). After modeling, the rats were randomly divided into sham group, model group, baicalin group, progesterone group and baicalin+mifepristone group. After 24 hours of modeling, score different time points of nerve function (6th hour,1st day,5th day and 10th day after modeling) by referring to Bederson method; The rats forehand grip on different time points of different groups was tested by holding power meter.Cerebral infarction volume fraction at the 10th day was determined by TTC staining.The baicalin normal group and baicalin group were given intraperitoneal injection of baicalin solution, the normal group, model group and sham group were given normal saline of the same quantity. Progesterone group was given intramuscular injection of progesterone. Baicalin+mifepristone group was given intraperitoneal injection of baicalin solution and intragastric administration of mifepristone solution. Serum was taken from the rats at the 10th day and the progesterone and related hormones levels of PROG, LH and FSH in the serum of every groups were measured by ELISA.Results1. To clarify the protective effect of baicalin on ischemic brain injury by the evaluation of neurological function in rats and analyse rat forelimb holding power rate and cerebral infarction volume percent. (1)Neural deficit scores:Compared with sham group, neurological function of modeling group rats were damaged after modeling 6h. There was no difference on nerve function score of modeling groups and every group of neurologic deficits are increased at the 1st day after modeling. Compared with model group, rat neural function in baicalin group and progesterone group at the 5th day trend to recover (P>0.05; P>0.05), and show more significantly improved at the 10th day (P<0.05; P<0.05). (2) Change rate of rat forelimb holding power:Compared with sham group,change rate of rat forelimb holding power was the most stongly reduced (P< 0.001) after modeling. Both baicalin group and progesterone group showed significantly improvement of rat forelimb holding power change rate at the 5th day(P <0.01; P<0.01),and high significantly improved the rate at the 10th day(P< 0.001; P< 0.001).(3) The result of TTC staining show that white infarction was not seen in rats brain slices of sham group.Other groups including model group, baicalin group, progesterone group and baicalin+mifepristone group all appeared different degrees of white infarcts. Compared with model group, cerebral infarction volume fraction of baicalin group and progesterone group decreased significantly (P<0.01; P<0.05), the fraction of baicalin+mifepristone group slightly decreased(P>0.05). There was no difference on the above curative effect between the baicalin group and progesterone group(P>0.05). The results showed that of the protective effect of baicalin on ischemic brain injury.2.To certify the regulating effect of baicalin on serum progesterone and correlated hormones level in normal and ischemic brain injury rats by using ELISA method to detect serum progesterone and related hormone levels. The results of serum hormone levels by ELISA method:rat serum was drawned and tested for hormone level at 10th day. Compared with normal group, progesterone level of the normal baicalin group has a tendency to increase(P>0.05). The level of luteinizing hormone (LH) decreased to some extent(P>0.05). Follicle stimulating hormone (FSH) levels increased slightly(P>0.05). Compared with model group, progesterone (PROG) level in baicalin group increased significantly(P< 0.05).Luteinizing hormone (LH) level decreased to some extent(P>0.05). Follicle stimulating hormone (FSH) level increased slightly (P>0.05). Progesterone (PROG), luteinizing hormone (LH) and follicle-stimulating hormone (FSH) level of baicalin+mifepristone group increased slightly(P> 0.05,P> 0.05,P> 0.05).This result prompt that baicalin can increase the normal group and model group serum progesterone level. Compared with normal group, model group progesterone level increased slightly, which may belong to cerebral ischemia model rats increased responses to stress, and LH and FSH levels have no obvious change.3. To explore the possible mechanism of the protective effect of baicalin on ischemic brain injury by comparing rats nerve function score, rat forelimb holding power rate, cerebral infarction volume percentage, the serum PROG and related hormone levels between the baicalin group and baicalin+mifepristone group.(1) Neural deficit scores:Compared with sham group, neurological function of modeling group rats were damaged, every group of neurologic deficits are increased at the 1st day after modeling. Compared with model group, rat neural function in baicalin group at the 5th day trend to recover (P> 0.05,and show more significantly improved at the 10th day(P<0.05). Compared with the baicalin group, baicalin+mifepristone group showed that the treatment effect was more significantly inhibited at the 10th day(P< 0.05). (2) Change rate of rat forelimb holding power:Compared with sham group, change rate of rat forelimb holding power was the most stongly reduced(P< 0.001) after modeling. Baicalin group showed significantly improvement of change rate rat forelimb holding power at the 5th day (P<0.01), and high significantly improved the rate at the 10th day(P<0.001). Compared with the baicalin group, baicalin+mifepristone group showed that the treatment effect was more significantly inhibited both at the 5th day and at the 10th day(P< 0.05). (3) The result of TTC staining show that white infarction was not seen in rats brain slices of sham group. Other groups including model group, baicalin group and baicalin+mifepristone group all appeared different degrees of white infarcts. Compared with model group, cerebral infarction volume fraction of baicalin group decreased significantly (P<0.01), the fraction of baicalin+mifepristone group slightly decreased(P>0.05). Compared with the baicalin group, the cerebral infarction volume fraction of baicalin+mifepristone group was higher than that of baicalin group(P>0.05). (4) The results of serum hormone level by ELIS A method:rat serum was drawned and tested for hormone level at 10th day. Compared with normal group, progesterone level of the normal baicalin group has a tendency to increase(P>0.05). The level of luteinizing hormone (LH) decreased to some extent(P>0.05). Follicle stimulating hormone (FSH) levels increased slightly(P>0.05). Compared with model group, progesterone (PROG) level in baicalin group increased significantly(P< 0.05).Luteinizing hormone (LH) level decreased to some extent(P>0.05).Follicle stimulating hormone (FSH) level increased slightly (P>0.05).Progesterone (PROG),luteinizing hormone (LH) and follicle-stimulating hormone (FSH) level of baicalin+mifepristone group increased slightly(P> 0.05,P> 0.05,P> 0.05). It prompt that baicalin can increase model rats serum progesterone leve, progesterone levels in the body by cerebral ischemia rats significantly was inhibited after application of mifepristone on progesterone block. It remainders that baicalin may play a role of brain protection by increasing serum progesterone level.ConclusionsBaicalin gave play to brain protection on ischemic brain injury model rats and improved the progesterone level of model rats.Baicalin's neurologic protection has been significantly suppressed and serum progesterone level significantly decreased after application of mifepristone on progesterone block.It remainders that baicalin may play a role of brain protection by increasing serum progesterone level. |
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