| Objective:This research revolved around basic pathogenesis of DKD. The analysis of multi-center clinical syndrome data was to explore the basic pathogenesis of diabetic kidney disease (DKD).Setting up items of scale for diagnosis of DKD qi deficiency and blood stasis syndrome was to summarize diagnosis elements of the most common syndrome,as well as clinical characteristics of basic pathogenesis.We also performed an animal experiment to examine the curative effect and mechanism of treatment according to the basic pathogenesis.Finally,we hope the results of this sutdy can provide the reference to the therapy of DKD in clinic.Methods:Study one:We retrospectively analysed multi-center clinical research syndrome data of DKD patients in stage three and four.Frequency method was uesd to analyse distribution of common syndrome elements of DKD patients in stage three, normal renal function of stage four, compensatory stage of renal insufficiency of stage four,as well as decompensated period of renal insufficiency. We also compared changes of common syndrome elements in different stages. Repeated measurement variance analysis was used to analyse the evolution of common syndrome elements in patients with follow-up of 24 months. And single factor variance analysis was used to compare difference of common syndrome elements integral at into the group, followed up for 12 months and 24 months.Study two:This research was based on the scale theory and referred production method of scale for syndromes diagnosis of Chinese medicine.Literature research was used to establish the item pool of scale for diagnosis of DKD qi deficiency and blood stasis syndrome. Delphi method was applied to filtrate items. Then,we established clinical questionnaire and surveyed 100 patients with DKD. Cronbach’s a and factor analysis were used to filtrate items,and evaluate the internal consistency reliability and structural validity of scale at the same time.Study three:23 GK rats were fed with high caloric and fat diet for 4 weeks.Then they were randomly divided into sham-operation group, model group, tonifying qi and promoting blood circulation group,as well as tonifying qi group.The model group, tonifying qi and promoting blood circulation group,and tonifying qi group were cut unilateral renal to establish diabetic glomerulopathy rat model. Sham-operation group and control group were filled the stomach with distilled water.Tonifying qi and promoting blood circulation group was filled the stomach with yi qi huo xue fang (radix astragali, winged euonymus twig) whose dosage was 10 times as the dosage of the adult.Tonifying qi group was filled the stomach with yi qi fang (radix astragali) whose dosage was also 10 times as the dosage of the adult.All groups were filled for 8 weeks.24 hour urine protein quantity of each group was tested at before gavage, after three weeks, six weeks and eight weeks. Then,we tested serum creatinine of all rats, observed renal pathological changes and the expression of AGE and TGF-β in kidney tissues by immunohistochemistry,western blot, real-time quantitative PCR,.Results:study one:This study included 544 cases of DKD patients,315 male,229 female,154 cases in DKD stage three,390 cases in DKD stage four.The distribution of syndrome elements in stage three was qi deficiency syndrome (71.7%), yin deficiency syndrome (66.4%), yang deficiency syndrome (50.1%),blood stasis syndrome (48.0%), phlegm dampness syndromes (45.4%), wet muddy syndrome (25.7%), blood deficiency syndrome (19.1%). The distribution of syndrome elements in normal renal function of stage four was qi deficiency syndrome (71.4%), yin deficiency syndrome (59.8%), yang deficiency syndrome (58.0%),blood stasis syndrome (51.8%), phlegm dampness syndromes (43.8%), blood deficiency syndrome (20.5%),wet muddy syndrome (8.0%).The distribution of syndrome elements in compensatory stage of renal insufficiency of stage four was qi deficiency syndrome (77.9%), yang deficiency syndrome (70.8%), blood stasis syndrome (66.2%), phlegm dampness syndromes (54.9%), yin deficiency syndrome (52.8%), wet muddy syndrome (29.7%), blood deficiency syndrome (28.2%).The distribution of syndrome elements in decompensated period of renal insufficiency of stage four was yang deficiency syndrome (80.0%), qi deficiency syndrome (71.8%), blood stasis syndrome(62.4%),yin deficiency syndrome (52.9%), phlegm dampness syndromes (48.2%), blood deficiency syndrome (32.9%), wet muddy syndromes (31.8%).Qi deficiency syndrome was the most common syndrome elements in different stages. Blood stasis syndrome was the most common syndrome elements of excessive pathogen. Proportion of yang deficiency syndrome, blood deficiency syndrome, and wet muddy syndrome proportion were increased gradually with the progress of the disease.Proportion of yin deficiency syndrome was gradually reduced.Phlegm dampness syndrome had no obvious change.The result of repeated measurement variance analysis for followed-up 287 patients showed that:different syndrome element integral was different from others at different time points.Each syndrome element integral was different at different time points. The integral of qi deficiency syndrome was higher than the others. The integral of blood stasis syndrome was similar to phlegm dampness syndrome.The integral of blood deficiency syndrome and wet muddy syndrome was relatively low.The results of pairwise comparison of common syndrome element integral showed that there was no difference between qi deficiency syndrome and yin deficiency syndrome at into groups and followed up for 12 months. At 24 months of follow-up, qi deficiency syndromes integral was higher than that of yin deficiency. Yang deficiency syndromes integral was lower than qi deficiency syndrome and yin deficiency syndrome at into the group. But at 12 months follow-up, yang deficiency syndromes integral was higher than yin deficiency syndrome.The integral of phlegm damp syndrome and blood stasis syndrome had no difference at full process of follow-up,as well as blood deficiency syndrome and wet muddy syndrome.The results of repeated measurement variance analysis for stage three and four were same as the followed-up 287 patients. The integral of yin deficiency syndrome was the highest in stage three, but had the downtrend.Qi deficiency syndromes integral was similar to yin deficiency syndrome. The integral of qi deficiency syndrome was the highest in stage four at into the group.But yang deficiency syndromes integral was similar to it of qi deficiency syndrome at 12 months follow-up. At 24 months of follow-up, yang deficiency syndromes integral became the highest, yin deficiency syndrome declined significantly. The results of pairwise comparison of common syndrome element integral in stage three showed that qi deficiency syndrome and yin deficiency syndrome integral were indifference, as well as phlegm damp syndrome and blood stasis syndrome. Qi deficiency syndrome, yin deficiency syndrome and yang deficiency syndrome had no difference at followed up for 12 months and 24 months. There were no difference between qi deficiency syndrome, yin deficiency syndrome and yang deficiency syndrome at into groups in stage four.At 12 months follow-up,yang deficiency syndrome was higher than yin deficiency syndrome. The change trend and distribution of all elements integral of control group were the same as foregoing results.Study two:through document retrieval of database,we collected 111 papers of DKD qi deficiency and blood stasis.Ruled out 72 papers which only mentioned the name of the qi deficiency and blood stasis syndrome, but did not mention corresponding symptoms.At last we extracted 40 common symptoms of deficiency syndrome and 29 symptoms of blood stasis.and established item pool.Setted up the first round questionnaire for experts according to item pool.In the first round,we consulted 20 experts from all over the country.The recovery rate was 100%. In the first round, the highest authority coefficient of experts was 0.80, the lowest was 0.65, the mean of it was 0.76.The coordination coefficient of expert was 0.28, chi-square was 314.03.Calculated mean and variation coefficient of each item.Deleted item whose the mean<2 and the variation coefficient >0.50. Finally,we deleted 25 items and reserved 32 items.In the second round,we also consulted 20 experts.The recovery rate was 95%.In the second round, the highest authority coefficient of experts was 0.80, the lowest was 0.65, the mean of it was 0.75. The coordination coefficient of expert was 0.20, chi-square was 104.44.At last,we deleted 4 item and reserved 32 items. According to the results of the two rounds of expert questionnaire,we setted up clinical questionnaire.100 patients were surveyed, including 29 patients in DKD stage three,44 patients in DKD stage four, and 27 patients in stage five,67 male,33 female.The Cronbach’s a of questionnaire was 0.70.Removed any item, the Cronbach’s a of entire questionnaire changed little, so kept all items. Factor analysis showed all items communality >0.50, the cumulative contribution of the common factor was 64.363%.After rotation, each item had the highest component matrix in one common factor. The rotated component matrix of each item were almost greater than 0.4. So all items retained. Finally,we established the item of diagnosis scale for DKD qi deficiency and blood stasis syndrome.Study three:after 1 week of establishing rat model,24 hour urine protein quantity of the model group was greater than 30mg/L,as well as tonifying qi and promoting blood circulation group and tonifying qi group.9 weeks after establishing rat model, the serum creatinine of model group rats were significantly higher than the control group. Renal pathological changes of each group conformed to DG. Before filled the stomach,24 hour urine protein quantitative of all groups had no difference. As time went on, urine protein was gradually increased of the model group and control group.24 hour urine protein quantitative of tonifying qi and promoting blood circulation group was same as tonifying qi group.They all significantly declined at 3 weeks after filled the stomach, and lower than that of model group and control group.But they all rised at 6 weeks.The mean of serum creatinine of tonifying qi and promoting blood circulation group was similar to the sham-operation group,which was also lower than the model group and tonifying qi group. HE staining results showed that pathological changes of tonifying qi and promoting blood circulation group was lighter than other groups. Immunohistochemical results showed the expression of AGE and TGF-β in kidney tissues were lower than control group and model group,as well as Western Blot. And real-time quantitative PCR results showed the expression of AGEmRNA and TGF-βmRNA in kidney tissues were lower than model group.Conclusion:qi deficiency and blood stasis is the basic pathogenesis of DKD.Qi deficiency and blood stasis syndrome is the most common clinical syndrome of DKD. According to production method of scale, we set up the item of diagnosis scale for DKD qi deficiency and blood stasis syndrome.Summarize the clinical characteristics of pathogenesis of qi deficiency and blood stasis.Basing on qi deficiency and blood stasis pathogenesis of DKD,the treatment of tonifying qi and promoting blood circulation can effectively treat DN, and its mechanism maybe reduce the expression of AGE and TGF-β. |
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