The Mechanism Study Of The Protective And Therapeutic Effect Of Semen Sojae Praeparatum Extract On Macroangiopathy In T2dm Rats

Objective: Macroangiopathy, the most frequent chronic complication in type 2 diabetes mellitus (T2DM), is the major factor to cause disability and death. Inflammatory reaction, oxidative stress which induced by hyperglycemia and hyperinsulinism, and the adjustment disorder of NO/NOS system are generally accepted as the major risk factors in its pathogenesis. To study the mechanism of the protective and therapeutic effect of Semen Sojae Praeparatum (SSP) on macroangiopathy in T2DM rats on both global and molecular level, our research investigated the alleosies of relative serum indicators, and the expression of nitric oxide synthase (NOS) mRNA in aorta of T2DM rats which induced by high glucose and fat diet in coordination with intraperitonal injection of small dose streptozotocin (STZ).Methods: To set up the T2DM model, fifty male Wistar rats (180-200g) were fed with high glucose and fat diet and injected with small dose STZ (25mg/kg) intraperitonally. To select forty rats whose blood glucose value>7.7mmol/L, and divided them into four groups at random: T2DM model group, low density group of SSP (10g crude drug/kg?d) , high density group of SSP (20g crude drug/kg?d) and rosiglitazone (RSG) group (1.8mg/kg?d), each group had ten rats. Besides, took ten rats as the normal control group. Normal control group and T2DM group were given distilled water orally, and the other groups were given respective drugs for eight weeks. The capacity that given to rats was 10ml/kg. The rats were decapitated to get the blood, and centrifugate the blood to get the serum eight weeks later. Measured the fast blood glucose (FBG), fast insulin (FIN) and superoxide dismutase (SOD), malondialdehyde (MDA), tumor necrosis factor-α(TNF-α), 6-ketonic-prostaglandin F1σ(6-Keto-PGF1σ), Thromboxane2 (TXB2), endothelin-1 (ET-1), nitric oxide (NO) in the blood serum. Scissored the rats' aorta quickly, and cut down about 1cm aorta from the superior extremity, and fixed it by 10% formaldehyde, then stained it by hematoxylin and eosin (HE) to make pathological sections. The residuary aorta was preserved in -80℃freezer. Extracted total RNA from the residuary aorta by Trizol, detected the expression of iNOS, eNOS mRNA by RT-PCR. All the datas were analysed with SPSS 11.5 and expressed with mean standard deviation.Results:1 The effect of SSP on FBG, FIN and ISI in T2DM ratsCompared with the NC group, the level of FBG in the T2DM group was obviously higher (p<0.01). Compared with the T2DM group, the level of FBG in the medication administration groups were all obviously lower (p<0.01 or p<0.05).The level of FIN and the value of ISI in T2DM group were obviously lower than those in the NC group (p<0.01). Compared with the T2DM group, the level of FIN and the value of ISI in the medication administration groups were all obviously higher (p<0.01).2 The effect of SSP on serum SOD and MDA in T2DMThe level of SOD in T2DM group was obviously lower than it in the NC group (p<0.01), the level of MDA in T2DM group was obviously higher than it in the NC group (p<0.01). Compared with the T2DM group, the level of SOD in the medication administration groups were all obviously higher (p<0.01 or p<0.05), the level of MDA in the medication administration groups were all obviously lower (p<0.01 or p<0.05).3 The effect of SSP on serum TNF-αin T2DM ratsCompared with the NC group, the level of TNF-αin the T2DM group was obviously higher (p<0.01). The level of TNF-αin the medication administration groups were all obviously lower than it in the T2DM group (p<0.01).4 The effect of SSP on serum 6-Keto-PGF1σand TXB2 in T2DM ratsThe level of 6-Keto-PGF1σin T2DM group was obviously lower than it in the NC group (p<0.01), the level of TXB2 in T2DM group was obviously higher than it in the NC group (p<0.01), and the value of TXB2/6-Keto-PGF1σwas obviously higher than it in the NC group (p<0.01). Compared with the T2DM group, the level of 6-Keto-PGF1σin the medication administration groups were all obviously higher (p<0.01 or p<0.05), the level of TXB2 in the medication administration groups were obviously lower (p<0.01), the value of TXB2/6-Keto-PGF1σin the medication administration groups were all obviously lower (p<0.01), too.5 The effect of SSP on serum ET-1 in T2DM ratsCompared with the NC group, the level of ET-1 in the T2DM group was obviously higher (p<0.01). The level of ET-1 in the medication administration groups were all obviously lower than it in the T2DM group (p<0.01).6 The effect of SSP on serum NO in T2DM ratsCompared with the NC group, the level of NO in the T2DM group was obviously higher (p<0.01). The level of NO in the medication administration groups were all obviously lower than it in the T2DM group (p<0.01).7 The effect of SSP on the appearance of aorta in T2DM ratsThe appearance of the aorta in NC group was: the intima of the aorta was intact and smooth; the tunic media had a clear courser, and no hyperplasia. The appearance of the aorta in T2DM group was: the intima of the aorta was coarse and obviously impaired, many inflammatory cells adhered in it; the tunic media was obviously hyperplastic, and the number of the cells was increased, some of the cell nucleus were hypertrophic. The appearance of the aorta in SSPL group was: the intima of the aorta was coarse and impaired, many inflammatory cells adhered in it; the tunic media obviously hyperplastic, the number of the cells in the superficial layer of the tunic media was increased. The appearance of the aorta in SSPH group was: compared with the T2DM group, the intima of the aorta was smooth, majority of the endothelial cells could be seen, part of the tunic intima was coarse, the number of cell nucleus was increased; the hyperplasia of the smooth muscles in tunic media was more lightly. The appearance of the aorta in RSG group was: compared with the T2DM group, the intima of the aorta was obviously smooth, the endothelium was intact, part of the intima was a bit hyperplastic; the hyperplasia of the smooth muscles in tunic media was more lightly.8 The effect of SSP on the expression of iNOS and eNOS mRNA in the aorta of T2DM ratsThe expression of iNOS mRNA in T2DM group was obviously higher than it in the NC group (p<0.01). Compared with the T2DM group, the expression of iNOS mRNA in the medication administration groups were all obviously lower (p<0.01).There was no notable difference of the eNOS mRNA expression among all the groups (p>0.05).Conclusions: SSP can decrease the FBG and FIN in the T2DM rats induced by high glucose and fat diet in coordination with intraperitonal injection of small dose STZ, and protect the aorta of it. One of the possible mechanism is promoting the anti-oxidation ability of T2DM rats to relieve the damage of oxidative stress. The other possible mechanism is reducing the release of the inflammatory factor TNF-αto relieve the damage of inflammatory reaction.The third possible mechanism is accommodating the NO/NOS system and the release of endothelial active factors ET-1 and TXA2, PGI2 to relieve the damage in endothelium.