| ObjectiveDiabetes mellitus induced erectile dysfunction(DIED)is a common complication of diabetes mellitus(DM).The incidence is high,which seriously affects the quality of life of patients and even the harmony of family and society.Due to a series of complex blood vessels and neuropathy caused by DM itself,DIED is more difficult to treat than common erectile dysfunction(ED)and belongs to refractory ED.Western medicine can effectively improve the erectile function of DIED patients,but it does not improve the patient’s systemic accompanying symptoms.Traditional Chinese medicine can make up for the deficiency of western medicine,and its position in DIED research is increasingly prominent.Huoxue Tongluo Qiwei Decoction is the experience of Professor Li Haisong of Dongzhimen Hospital.It has the function of promoting blood circulation and removing blood stasis,and the decoction has been proven to improve the erectile function and systemic symptoms of ED patients with blood stasis and kidney deficiency.The purpose of this study was to further evaluate the clinical efficacy of Huoxue Tongluo Qiwei Decoction on the blood stasis and kidney deficiency type DIED on the basis of the western medicine tadalafil and to observe the endothelial damage and platelet of DIED rat model by promoting Huoxue Tongluo Qiwei Decoction.The effects of abnormal activation,protein kinase C-β(PKCβ)signaling pathway-related molecular expression,and further explore its mechanism of action.MethodsClinical study:A randomized controlled trial was conducted to enroll 82 patients with ED induced by type 2DM who were eligible for blood stasis and kidney deficiency,and were randomly assigned to a treatment group and a control group of 41 patients.The control group received a small dose of tadalafil tablets(5mg)at 15:00 daily,the treatment group was given a daily dose of Huoxue Tongluo Qiwei Decoction Granules on the basis of the control group,2 times a day in the morning and evening,and the treatment course was 4 weeks.Detailed records of each patient’s general information,symptoms,signs,treatment history and combined diseases,international index of erectile function(IIEF-5)scores and TCM syndrome scores before and after treatment,and comparison,detailed records Adverse reactions that occur during treatment.Experiments:1.80 male Sprague-Dawley rats were randomly divided into a blank control group(K group,n=10)and prefabricated modules(n=70).Group K was intraperitoneally injected with sodium citrate buffer;70 precast rats were intraperitoneally injected with streptozotocin(STZ)solution in the right lower quadrant at 55 mg/kg.Blood samples were taken from the tail vein at 72h and 1 week after injection.The blood glucose was higer than 16.7mmol/L after 1 week,and the urine volume was increased.The diet was determined to be the DM rat model.Eight weeks after the injection,63 of the remaining DM rats were injected with apomorphine(APO)for screening,and a total of 45 DIED rats were obtained.40 rats were randomly selected from and randomly divided into model group(M group),LY333531 inhibitor group(L group),Huoxue Tongluo Qiwei decoction low dose group(D group),Huoxue Tongluo Qiwei decoction high dose Group(G group),all rats were treated with deionized water or corresponding drugs for 8 weeks;2.Observed the body weight,blood glucose level,erection times,vascular endothelial injury markers of each group after drug intervention Lectin-like oxidized low density protein receptor-1(LOX-1),soluble E-selectin(sE-selectin),soluble intercellular adhesion molecule-1(sICAM-1),endothelial cell microparticles(EMPs),Oxidative stress markers superoxide dismutase(SOD),malondialdehyde(MDA),platelet activation marker P-selectin(CD62P)levels,and pathomorphology of corpus cavernosum,ultrastructure of corpus cavernosum vascular endothelial cells 3.Western blot and Real-Time PCR were used to detect the peptides involved in the PKCβ signaling pathway in rat corpus cavernosum,such as Diacylglycerol(DAG),PKCβ,Nuclear factor kappa-B(NF-κB),and cells.Expression of protein and mRNA of Intercellular adhesion molecule-1(ICAM-1).ResultsClinical study:During the observation period,1 patient in the treatment group and 2 patients in the control group fell off,and 79 patients eventually ended.The treatment group was cured in 5 cases,markedly effective in 18 cases,effective in 7 cases,ineffective in 10 cases,the total effective rate was 75.0%;in the control group,2 cases were cured,10 cases were markedly effective,6 cases were effective,21 cases were ineffective,and the total effective rate was 46.1%.The treatment group was better than the control group.The improvement of IIEF-5 score in the treatment group was more significant than that in the control group(P<0.05).In terms of TCM syndrome scores,both the treatment group and the control group were able to significantly improve the TCM syndrome total score and TCM main symptom scores in patients with DIED(P<0.05),and the treatment group was better(P<0.05).The two groups in terms of secondary symptoms of Chinese medicine,thoracic tingling,abdominal detention,weak waist and knees,chills and cold limbs,god fatigue,and tinnitus were statistically different(P<0.05),and after treatment,there is no significant difference in perineal swell and scrotal pain(P>0.05).There was 1 headache in the treatment group,1 facial flushing in the control group,and 1 mild back pain.The above adverse reactions were mild and could be relieved by themselves without special treatment.Experiments:The body weight of rats in DIED group was significantly different from that in group K(P<0.05).There was no significant difference between the body weight of each DIED group(P>0.05);DIED group rats The blood glucose was significantly higher than that of the K group(P<0.05).The blood glucose levels of the G and D groups were relatively better than those of the M and L groups,but there was no statistical difference in the blood glucose levels between the DIED groups(P>0.05).2.induced by APO After that,the number of induced erections in group M was 0,which was significantly different from that in group K(P<0.05).After drug intervention,the number of erections in rats in group L,D,and G was different from that in group M(P<0.05),G group was better than L and D groups,but there was no significant difference between the groups(P>0.05).3.Compared with group K,serum LX-1,sE-of group M rats.The levels of selectin and sICAM-1 were significantly increased(P<0.05).After drug intervention,the serum levels of LOX-1,sE-selectin and sICAM-1 were significantly higher in the L,D and G groups than in the M group.Decreased(P<0.05).The improvement of LOX-1,sE-selectin and sICAM-1 levels in group G and group D was better than that in group L,but there was no significant difference between groups(P>0.05).4.with K Compared with the group,the serum MDA level of the M group was significantly increased.The level of SOD was significantly lower(P<0.05).After drug intervention,compared with group M,the serum MDA of rats in group L,D and G decreased significantly,and the level of SOD increased significantly(P<0.05).Groups G and L were MDA and G group were relatively better in SOD improvement,but there was no significant difference in MDA and SOD levels between the groups(P>0.05).5.Compared with group K,EMPs levels were significantly higher in group M(P<0.05).After drug intervention,the levels of EMPs in the L,D,and G groups were significantly lower than those in the M group(P<0.05),and the G group was significantly decreased compared with the L and D groups.There was no significant difference in EMPs between the groups(P>0.05).6.Compared with group K,CD62P level was significantly increased in group M(P<0.05).After drug intervention,compared with group M,CD62P levels in rats in group L,D and G decreased significantly(P<0.05),G and D groups were more obvious than L group,and there was no significant difference in CD62P levels between the groups(P>0.05).7.Under light microscope,compared with K group,group M corpus cavernosum sinusoidal and small The number of blood vessels was significantly reduced,the distribution of sinusoids was disordered,the density of endothelial cells and the density of smooth muscle cells were significantly decreased,and the number of collagen fibers was significantly increased.After drug intervention,the distribution of sinusoids in rats of group L,D and G was higher than that of M.Relatively regular,the density of endothelial cells increased,the number of collagen fibers decreased,and red blood cells were partially visible in some blood sinus spaces.Under electron microscope,M group showed more internal rupture of sinusoidal endothelium,narrowing of sinusoidal cavity,damage of endothelial cell plasma membrane,swelling,obvious reduction of organelles,swelling of mitochondria,expansion of endoplasmic reticulum,and even no organelles.The nuclear chromatin edge is concentrated.After drug intervention,although a certain degree of endothelial proliferation was observed in the L,D,and G groups,it was also observed that the nucleus of the M group was relatively intact,and the organelles such as mitochondria,Golgi apparatus,and endoplasmic reticulum increased significantly,or the organelles were basically Normally improved significantly.8.In Western blot,the protein expression levels of DAG,PKCβ,NF-κB and ICAM-1 were significantly increased in M group compared with K group.After administration,DAG in group L,D and G,The expressions of PKCβ,NF-κB and ICAM-1 were significantly down-regulated compared with M group(P<0.05).Among them,the D and G groups improved in the expression of PKCβ,NF-κB and ICAM-1 in DAG and G groups.Relatively more obvious,but there was no significant difference between the three groups of L,D and G(P>0.05).9.In the Real-Time PCR test,the M group rats were compared with the K group rats in DAG,PKCβ,NF-κB,ICAM.The mRNA expression level of-1 was significantly increased.After administration,the expressions of DAG,PKCβ,NF-κB and ICAM-1mRNA in L,D and G groups were significantly lower than those in M group(P<0.05).The numerical improvement of DAG,PKCβ and ICAM-1 expression in group G was relatively more obvious,but there was no significant difference between L,D and G groups(P>0.05).Conclusion1.Huoxue Tongluo Qiwei Decoction combined with low-dose tadalafil can safely and effectively improve the erectile function of patients with blood-stagnation and kidney-deficiency of DIED.The clinical efficacy is better than the low-dose tadalafil alone.2.Huoxue Tongluo Qiwei Decoction can improve the erectile function of DIED rats,and its mechanism may relate to reducing endothelial damage,reducing oxidative stress damage,improving the body’s antioxidant capacity,inhibiting platelet abnormal activation,and repairing the structure of corpus cavernosum vascular endothelial cells of DIED rats to some degree.3.The mechanism of DIED may be related to vascular endothelial injury and abnormal platelet activation caused by activation of PKCβ signaling pathway;Huoxue Tongluo Qiwei Decoction protects endothelial function and anti-platelet activation by inhibiting the expression of PKCβ signaling pathway-related molecules DAG,PKCβ,NF-κB and ICAM-1 protein and mRNA in DIED rats. |
