摘要
近年研究提示,炎症和免疫是糖尿病肾病的重要发病机制,本研究通过实验研究和临床研究两方面,分别观察NF-κB炎症信号通路,免疫受体Fcγ RIIB/C蛋白在糖尿病肾病发生发展过程中的作用。
实验研究：黄芩素在高糖诱导人近端肾小管上皮细胞损伤中的作用和机制研究
NF-κB作为一个炎症信号通路的关键因子,其介导的炎症反应在糖尿病肾病的病理发展过程中起着十分重要的作用,但其在糖尿病近端肾小管上皮细胞损伤中的作用尚不明确。本实验通过体外研究发现：NF-κB信号激活参与了高糖诱导的人近端肾小管上皮细胞(HK-2)损伤；中药黄芩的单体成分—黄芩素,可能通过抑制NF-κB通路激活,减轻高糖诱导的肾脏细胞炎症状态。
目的：
1.观察高糖对HK-2细胞糖毒性作用及相关信号通路。
2.黄芩素干预高糖诱导HK-2细胞损伤的作用及机制研究。
方法：
不同糖浓度(5.5mM-60mM)和不同甘露醇浓度(30mM,45mM,分别对应各高糖渗透压)孵育HK-2细胞24h,观察细胞活力变化。不同糖浓度(5.5mM-45mM)刺激HK-2细胞24h,观察IκBα-和p65蛋白表达规律。不同剂量的黄芩素(25-200uM)处理未加刺激的HK-2细胞36h,观察细胞活力变化。选择45mM浓度的葡萄糖刺激HK-2细胞24h,将NF-κB特异抑制剂PDTC作为阳性对照,观察不同剂量黄芩素(25-100uM)对高糖诱导的HK-2细胞NF-KB通路激活及其下游炎症因子和细胞外基质的影响。MTT比色法检测细胞活力。Western blot法检测NF-κB信号通路关键蛋白(IKBa、p65、 p-p65、IKKa),及其下游炎症分子单核细胞趋化蛋白-1(MCP-1)、细胞间黏附分子-1(ICAM-1)和细胞外基质重要成分IV型胶原蛋白(Collagen Ⅳ)表达。
结果：
1.高糖诱导的HK-2细胞糖毒性损伤
不同糖浓度(5.5mM、15mM、30mM、45mM、60mM)孵育HK-2细胞24h后,MTT法检测细胞活力。5.5maM浓度的葡萄糖视为正常糖对照组。结果发现不同糖浓度呈浓度依赖性对HK-2细胞活力存在抑制作用,与正常糖对照组比较,除15mM浓度的葡萄糖外(P>0.05),30mM、45mM、60mM浓度的葡萄糖差异有显著性(P<0.05,P<0.01);30mM、45mM浓度的葡萄糖与60mM浓度的葡萄糖比较差异有显著性(P<0.05)；30mM与45mM浓度的葡萄糖比较差异无显著性(P>0.05)。不同甘露醇浓度(35mM,45mM,分别对应各高糖渗透压)孵育HK-2细胞24h后,结果发现与止常糖对照组比较,30mM.45mM浓度的甘露醇对HK-2细胞活力没有明显的抑制作用(P>0.05)；30mM与45maM浓度的甘露醇比较差异无显著性(P>0.05)。上述结果提示高糖诱导的HK-2细胞毒性损伤呈一定的浓度依赖关系,且与渗透压作用无明显关联。结合国内外相关研究及死亡细胞过多会影响实验结果的经验,本研究初步选择5.5mM-45mM浓度的葡萄糖进行下一步实验。
2.高糖诱导的HK-2细胞NF-κB通路激活的作用
Western blot结果发现：IκBa蛋白表达随着葡萄糖浓度的升高而逐渐降低,与正常糖对照组比较,30mM、45mM浓度的葡萄糖差异有显著性(P<0.05,P<0.01)；30mM与45mM浓度的葡萄糖比较差异无显著性(P>0.05)。p65蛋白随葡萄糖浓度的升高而表达上调,30maM、45mM浓度的葡萄糖与正常糖对照组比较差异有显著性(P<0.05,P<0.01)；30mM与45mM浓度的葡萄糖比较差异无显著性(P>0.05)。提示高糖刺激HK-2细胞下调IκBα蛋白表达和上调p65蛋白表达,呈一定的浓度依赖关系。因此,本研究选择45mM浓度的葡萄糖作为高糖刺激组,进行黄芩素干预实验研究。
3.黄芩素对正常培养的HK-2细胞活力的影响
MTT法实验结果所示：25uM、50uM、100uM剂量的黄芩素对未加刺激正常培养的HK-2细胞活力没有明显的抑制作用,与空白对照组比较差异无显著性(P>0.05);200uM的黄芩素对正常培养的HK-2细胞活力有明显的抑制作用,与空白对照组比较差异有显著性(P<0.05)。提示200uM剂量的黄芩素对HK-2细胞存在一定的细胞毒性。基于本实验结果,初步选择25uM、50uM、100uM剂量的黄芩素,分别设为黄芩素低剂量组、黄芩素中剂量组、黄芩素高剂量组进行下一步实验。
4.黄芩素对高糖诱导的HK-2细胞NF-κB通路激活的作用
1)黄芩素干预抑制高糖诱导的HK-2细胞IκBa蛋白表达下调
Western blot实验结果所示：100uM的NF-κB特异抑制剂PDTC对高糖诱导的HK-2细胞IκBa蛋白表达下调有明显的抑制作用,与高糖组比较有显著差异(P<0.01)；黄芩素可以剂量依赖性的抑制高糖诱导的HK-2细胞IKBa蛋白表达下调,其中黄芩素中、高剂量组与高糖组比较有显著差异(P<0.05,P<0.01),黄芩素低剂量组与高糖组比较无显著差异(P>0.05)。
2)黄芩素抑制高糖诱导的HK-2细胞p65活化
Western blot实验结果所示：高糖刺激HK-2细胞24h可显著上调p-p65/p65比值,与正常糖对照组比较有显著差异(P<0.01)；黄芩素中、高剂量组和PDTC组能抑制高糖诱导的HK-2细胞p-p65/p65比值上调,与高糖组比较差异有显著性(P<0.05,P<0.01)；黄芩素低剂量组与高糖组比较无显著差异(P>0.05)。提示高糖通过磷酸化作用活化p65促进其向胞核转位；黄芩素呈一定的剂量依赖关系阻断p65活化。
3)黄芩素干预抑制高糖诱导的HK-2细胞IKKa表达
Western blot实验结果所示：高糖刺激HK-2细胞24h可显著上调IKKa蛋白表达,与正常糖对照组比较有显著差异(P<0.01)；黄芩素中、高剂量组能下调高糖诱导的HK-2细胞IKKa蛋白表达,与高糖组比较有显著差异(P<0.05,P<0.01)。结合上述结果,提示黄芩素可能通过抑制IKKa介导的IκBα降解和p65磷酸化,抑制NF-κB通路激活。
5.黄芩素干预对高糖诱导的HK-2细胞炎症分子MCP-1、ICAM-1表达的影响
Western blot实验结果所示：高糖刺激24h可诱导HK-2细胞MCP-1、ICAM-1蛋白表达上调,与正常糖对照组比较有显著差异(P<0.01)；黄芩素中、高剂量组和PDTC组能下调高糖诱导的MCP-1、ICAM-1蛋白表达,与高糖组比较有显著差异(P<0.05,P<0.01)。
6.黄芩素干预对高糖诱导的HK-2细胞外基质成分Collagen IV表达的影响
Western blot实验结果所示：高糖刺激24h可诱导HK-2细胞Collagen IV蛋白表达上调,与正常糖对照组比较有显著差异(P<0.01)；黄芩素中、高剂量组和PDTC组能下调高糖诱导的Collagen IV蛋白表达,与高糖组比较有显著差异(P<0.05,P<0.01)。
结论：
1.高糖诱导的HK-2细胞毒性损伤呈一定的浓度依赖关系,且与渗透压作用无明显关联。高糖能够刺激HK-2细胞1KKα/IκBα/p65通路激活,并上调其下游炎症分子MCP-1、ICAM-1表达,及促进ECM成分Collagen IV堆积。
2.黄芩素可以通过抑制高糖诱导的HK-2细胞IKKα/IκBα/p65通路激活,下调其下游炎症分子MCP-1、ICAM-1表达,减少ECM成分Collagen IV堆积。提示黄芩素可以通过减轻肾脏细胞炎症来延缓糖尿病肾病的进展,且与抑制NF-κB炎症信号通路的激活密切相关。
临床研究：Fcγ RIIB/C与糖尿病肾病相关性及中医证型分析
在第一部分研究中,我们证实,炎症最关键的通路NF-kB的激活参与了DN的发生发展。以往的研究发现,促炎症免疫受体Fcγ Rs (Fc gamma receptors)可以刺激外周血中性粒细胞NF-kB通路激活,提示FcyRs与NF-κB密切相关。
Fcγ Rs是免疫球蛋白G(IgG)恒定区Fc的受体。最新的研究发现,激活型Fcγ Rs参与免疫复合物介导的糖尿病小鼠肾脏炎症过程,而抑制型Fcγ RIIB能够抑制激活型Fcγ Rs的功能；且C反应蛋白(CRP)参与Fcγ Rs和NF-κB的激活。因此,观察FcγRIIB/C在糖尿病肾病中的表达规律,分析其与超敏C反应蛋白(hs-CRP)等相关指标的关系,对糖尿病肾病的临床与实验研究可能具有一定的价值。
目的：
1.观察血清Fcγ RIIB/C在糖尿病肾病中的表达规律；
2.观察血清hs-CRP在糖尿病肾病中的表达规律;
3.分析Fcγ RIIB/C与hs-CRP等相关指标的关系；
4.分析糖尿病肾病中医证型分布特点；
5.分析糖尿病肾病中医证型与hs-CRP的关系。
方法：
研究对象为2012年6月至2013年1月我院内分泌科和肾病科住院的2型糖尿病(含糖尿病肾病)患者共53例。参照文献根据尿白蛋白/肌酐比值分为：①糖尿病正常蛋白尿组(DM)：尿白蛋白/肌酐比值<40mg/g,其中男8例,女10例,年龄55.44±14.51岁；②糖尿病微量蛋白尿组(DNl)：尿白蛋白／肌酐比值为40～300mg/g,其中男10例,女8例,年龄59.15±13.45岁；③糖尿病临床蛋白尿组(DN2):尿白蛋白/肌酐比值为>300mg/g,其中男8例,女9例,年龄62.60±8.73岁。另设健康对照组(NC)：10例,其中男4例,女6例,年龄52.00±13.58岁。记录临床基本资料,如年龄、身高、体重、病程、血压、糖化血红蛋白、总胆固醇、甘油三酯、低密度脂蛋白、高密度脂蛋白、血肌酐、胰岛素抵抗指数、超敏C反应蛋白等,并进行中医证型分析。酶联免疫吸附法检测血清Fcγ RIIB/C蛋白水平。
结果：
1.各组基本临床资料比较结果显示：DN2组患者年龄最大,与NC组比较有显著差别(P<0.05)。DN1组和DN2组患者病程明显延长,与DM组比较有显著差别(P<0.05)。DN1组总胆固醇最高,与DM组比较有显著差别(P<0.05)。NC组、DM组和DN2组低密度脂蛋白低于DN1组,差别具有显著性(P<0.05)。DM组、DN1组和DN2组胰岛素抵抗指数高于NC组,其中DM组和DN2组与NC组比较有显著(P<0.05)。血压、体重指数、甘油三酯、血肌酐、糖化血红蛋白、高密度脂蛋白各组间比较无显著差别(P>0.05)。
2.DM组(18.06±5.34)血清Fcγ RIIB/C浓度显著低于NC组(25.28±8.96)、DN1组(26.75±9.15)和DN2组(24.09±8.39),差别具有显著性(P<0.05,P<0.0I)。DN1和DN2组血Fcγ RIIB/C浓度与NC组相近,差别无显著性(P>0.05)。
3.DM组(2.06±1.60)、DN1组(3.21±2.20)、DN2组(1.90±2.14)血清hs-CRP浓度显著高于NC组(0.86±0.94)(P<0.05,P<0.01)。DN1组血清hs-CRP浓度高于DM组和DN2组,但三组无显著差异(P>0.05)。
4.血清Fcγ RIIB/C仅与hs-CRP呈负相关(r=-0.276,P=0.048),与年龄、病程、血压、体重指数、总胆固醇、甘油三酯、高密度脂蛋白、低密度脂蛋白、血肌酐、胰岛素抵抗指数、尿白蛋白/肌酐比值均无明显相关。
5.本证证型中未见阴阳两虚证；阴虚燥热证、气阴两虚证、脾肾气虚证三个本证中,以气阴两虚证为主,显著高于阴虚燥热证、脾肾气虚证(P<0.01)。标证以血瘀证、湿热证为主,血瘀证显著多于湿热证(P<0.01)。
6.糖尿病肾病患者本证证型中,阴虚燥热证(2.40±1.06)、气阴两虚证(3.84±1.93)、脾肾气虚证(2.13±0.90)血清hs-CRP浓度显著高于健康对照组(0.86±0.94)(P<0.05,P<0.01),气阴两虚证血清hs-CRP浓度显著高于阴虚燥热、脾肾气虚组(P<0.05)。标证证型中,湿热证(4.28±2.12)、血瘀证(1.83±1.65)血清hs-CRP浓度与健康对照组比较有显著差异(P<0.05,P<0.01)；湿热证血清hs-CRP浓度显著高于血瘀证(P<0.05)。
结论：
1.糖尿病肾病患者血清Fcγ RIIB/C浓度没有显著变化,不能为糖尿病肾病诊断和治疗提供依据；
2.糖尿病肾病患者存在微炎症状态；
3.血清Fcγ RIIB/C与微炎症状态呈负相关；
4.气阴两虚,瘀血阻络兼夹湿热是糖尿病肾病早中期的基本病机；
5. hs-CRP水平与中医证型相关,在一定程度上为中医辨证分型提供客观依据。
The impact of NF-kB inflammatory signaling pathway and immune receptor Fcγ RIIB/C on the development and progression of diabetic nephropathy were observed respectively from experimental and clinical research.
Experimental research—The role of baicalein in high glucose-i nduced human renal proximal tubular epithelial cell(HK-2) injury
NF-kB-mediated inflammatory response, as a key factor of an inflammatory signaling pathway, plays a very important role in the pathological process of diabetic nephropathy, but the role of NF-kB in high glucose-induced human renal proximal tubule epithelial cell injury is not clear. In vitro study, we demonstrated that NF-kB activation took part in renal tubular epithelial cell injury induced by high glucose, and baicalein could inhibit NF-kB-mediated inflammatory response in high glucose (HG)-induced HK-2cell injury.
Objective:
1. To investigate the toxic effects of high glucose on human renal tubular epithelial cell line (HK-2) and related signaling pathway.
2. To investigate the role of baicalein in HG-induced HK-2cell injury.
Methods:
Different glucose concentrations (5.5mM-60mM) and mannitol concentrations (30mM, 45mM, respectively, corresponding to the osmotic pressure of the high glucose) stimulated HK-2cells for24h, cell viability was observed. Different glucose concentrations (5.5mM-45mM) stimulated HK-2cells for24h, the expression of IκBα and p65were observed.Baicalein (25-200uM) stimulated HK-2cells for36h with any treatment, cell viability was observed. HG (45mM) was selected to stimulate HK-2cells for24h, the NF-kappaB inhibitor PDTC was used as a positive control.To observe the inhibitory effect of different doses of baicalein (25-100uM) on NF-kappaB pathway activation and its downstream inflammatory molecules and extracellular matrix in HG-induced HK-2cells. Cell viability was detected by MTT assay. The key proteins of NF-kappaB signaling pathway (IκBa, p65, p-p65, IKKa), and its downstream inflammatory molecules (MCP-1, ICAM-1) and an important component of the extracellular matrix (Collagen IV) were measured by Western blot analysis.
Result:
1. The toxic effect of high glucose on HK-2cells
Different glucose concentrations (5.5mM,15mM,45mM,60mM) were incubated for HK-2for24h, cell viability was detected by MTT assay, normal glucose (5.5mM) as control group (NG). Found that the different glucose concentrations had an inhibitory effect on cell viability of HK-2cells in a dose-dependent manner.compared with NG, HG (30mM,45mM,60mM) had significant difference(P<0.05,P<0.01); compared with HG (60mM), HG(30mM,45mM) had significant difference (P<0.05);compared with HG(30mM), HG(45mM) had no significant difference (P>0.05). Mannitol concentrations (30mM,45mM, corresponding to the osmotic pressure of high glucose, respectively) were incubated for HK-2cells for24h. Found that HM(30mM,45mM) had no significant inhibitory effect on cell viability of HK-2cells compared with NG (P>0.05); compared with HM(30mM), HM(45mM) had no significant difference (P>0.05).These results suggested that the toxic effect of high glucose on HK-2cells was dose-dependant, which had no significant correlation with the osmotic pressure. According to domestic and foreign research and dead cells too much will affect the experimental results, glucose concentrations(55mM to45mM) were subjected to further analysis.
2. The role of NF-κB activation in HG-induced HK-2cell injury
Expression of IκBa and p65in HK-2cells following exposure to different glucose concentrations (5.5-45mM) for24h and were measured by Western blot. The results showed that hκBa protein expression decreased with increasing glucose concentration, compared with NG, HG(30mM,45mM) had significant difference (P<0.05, P<0.01) compared with HG(30mM),HG(45mM) had no significant difference (P>0.05).p65protein expression increased with increasing glucose concentration, compared with NG, HG(30mM,45mM) had significant difference (P<0.05,P<0.01); compared with HG(30mM), HG(45mM) had no significant difference (P>0.05).The results demonstrated that down-regulation of IκBα protein and up-regulation of p65protein in HG-induced HK-2cell injury was dose-dependant. Therefore, HG (45mM) was selected for future baicalein study.
3. The effect of baicalein on cell viability of HK-2cells without any treatment
MTT assay showed that baicalein (25uM,50uM,100uM) had no significant inhibitory effect on cell viability of HK-2cells without any treatment compared with control group (P>0.05).Baicalein (200uM) had an significant inhibitory effect on cell viability of HK-2cells without any treatment compared with control group (P<0.05).The results suggested that baicalein (200uM) had toxicity to HK-2cells. Based on the experimental results, baicalein (25uM,50uM,100uM) was selected initially for further study.
4. The effect of baicalein on NF-κB activation in HG-induced HK-2cells
1) The inhibitory effect of baicalein on down-regulation of IκBα protein in HG-induced HK-2cells.
Western blot assay showed that NF-κB specific inhibitor PDTC(100uM) had an significant inhibitory effect on down-regulation of IκBα protein compared with HG (P<0.01).Baicalein suppressed the down-regulation of IκBα protein in a dose-dependant manner, Baicalein(50uM,100uM) were significantly different compared with HG(P<0.05, P<0.01),Baicalein (25uM) showed no significant difference compared with HG (P>0.05)
2) The inhibitory effect of baicalein on p65activation in HG-induced HK-2cells.
Western blot assay showed that the p-p65/p65ratio was raised in HG-induced HK-2cells compared with NG (P<0.01); Baicalein(50uM,100uM) and PDTC(100uM) down-regulated the p-p65/p65ratio compared with HG (P<0.05,P<0.01); Baicalein (25uM) showed no significant difference compared with HG(P>0.05).The results implied that high glucose could activate p65to promote its translocation to nucleus through phosphorylation of p65; and baicalein inhibited p65activation in a dose-dependant manner.
3) Baicalein inhibited HG-induced protein expression of IKKa in HK-2cells.
Western blot assay showed HG treatment for24h markedly increased IKKa protein expression compared with NG (P<0.01);Baicalein(50uM,100uM) decreased IKKa protein expression compared with HG(P<0.05,P<0.01).The combination of these results implied that baicalein inhibited NF-κB pathway activation through IKKa-mediated IκBα degradation and p65phosphorylation in HG-induced HK-2cells.
5. The effect of baicalein on inflammatory molecules (MCP-1, ICAM-1) in HG-induced HK-2cells.
Western blot assay showed that HG treatment for24h markedly increased the MCP-1and ICAM-1protein expression compared with NG (P<0.01).Baicalein(50uM,100uM) and PDTC(100uM) decreased MCP-1and ICAM-1protein expression compared with HG (P<0.05, P<0.01)
6. The effect of baicalein on extracellular matrix component Collagen Ⅳ in HG-induced HK-2cells.
Western blot assay showed that HG treatment for24h markedly increased the Collagen Ⅳ protein expression compared with NG (P<0.01); Baicalein(50uM,100uM) and PDTC(100OuM) decreased Collagen IV protein expression compared with HG(P<0.05, P<0.01)
Conclusion:
1. The toxic effect of high glucose on HK-2cells was dose-dependant, which had no significant correlation with the osmotic pressure.IKKα/IκBα/p65signaling pathway was activated, its downstream inflammatory molecules (MCP-1, ICAM-1) were up-regulated, and extracellular matrix component Collagen Ⅳ was increased in HG-induced HK-2cell injury.
2. Baicalein could suppress IKKα/IκBα/p65signaling pathway, down-regulated the protein expression of MCP-1、ICAM-1and Collagen Ⅳ in HG-induced HK-2injury. The results suggested that NF-κB activation took part in renal tubular epithelial cell injury induced by high glucose, and baicalein could inhibit NF-κB-mediated inflammatory response in HG-induced HK-2cell injury.
Experimental research—the change of serum Fcγ RIIB/C in diabetic nephropathy and Analysis of TCM syndromes
In the first part of the study, we demonstrated that NF-kB signaling pathway involved in the development of DN. Previous studies had found that the proinflammatory immune receptors Fcγ Rs (Fc gamma receptors) could stimulate NF-kB activation in peripheral blood neutrophils, which implied that Fcγ Rs and NF-kB were closely related.
Fcγ Rs is a constant region of immunoglobulin G (IgG) Fc receptors. In the latest study, Virginia proposed that activating FcyRs is required for initiating immuncomplex-mediated inflammation in the diabetic kidney, but limited in its response by inhibitory Fcγ RIIB.In addition, C-reactive protein (CRP) involved in the Fcγ Rs and NF-kB activation. Therefore, it might be some valuable to observe the change of Fcγ RIIB/C in diabetic nephropathy and its correlation with related indicators.
Objective:
1. To observe the change of serum Fcγ RIIB/C in diabetic nephropathy;
2. To observe the change of serum hs-CRP in diabetic nephropathy;
3. To observe the correlation of Fcγ RIIB/C with hs-CRP and other related indicators;
4. To analyze syndrome distribution of traditional Chinese medicine (TCM);
5. To analyze the correlation of TCM syndromes with hs-CRP.
Methods:
53cases of patients were divided into three groups according to the urinary albumin/creatinine ratio:①diabetes with normal albuminuria group (DM), urinary albumin/creatinine ratio<40mg/g, including8males,10females, aged55.44±14.51years;②diabetes with microalbuminuria group (DN1), urinary albumin/creatinine ratio of40to300mg/g including10males and8females, aged59.15±13.45years;③diabetes with clinical albuminuria group (DN2),urinary albumin/creatinine ratio>300mg/g, including8males and9females, aged62.60±8.73years. In addition,10cases of healthy subjects as normal control group (NC), including4males and6females, aged52.00±13.58years.The basic clinical information, such as age, height, weight, blood pressure, HbAlc, hs-CRP,HOMA-IR,TC,TG,LDL,HDL,Scr and so on,were recorded. At the same time, TCM syndrome types were analyzed. Serum Fcγ RIIB/C level was measured by enzyme-linked immunosorbent assay.
Results:
1. DN2was the oldest compared with NC (P<0.05).The duration of DN2and DN2were longer compared with DM (P<0.05).TC increased in DN2compared with DM (P<0.05). LDL decreased in NC,DM,DN2compared with DN1(P<0.05).HOMA-IR increased in DM,DN2compared with NC (P<0.05).There was not any difference of blood pressure,BMI,TG, HDL,Scr,HbA1c among three diabetic groups and NC (P>0.05).
2. Serum Fey RIIB/C decreased in DM (18.06±5.34) compared with NC (25.28±8.96),DN1(26.75±9.15),DN2(24.09±8.39)(P<0.05, P<0.01),but there was no difference in NC,DN1,DN2(P>0.05).
3. Serum hs-CRP increased in DM (2.06±1.60),DN1(3.21±2.20),DN2(1.90±2.14) compared with NC (P<0.05, P<0.01),but there was no difference in DM,DN1,DN2.
4. Serum Fey RIIB/C had a weak negative correlation with Serum hs-CRP(r=-0.276, P=0.048). Serum Fcγ RIIB/C had no correlation with age, duration, blood pressure, BMI, TC, TG, LDL, HDL, Scr, HOMA-IR, urinary albumin/creatinine ratio.
5. In the four deficiency syndromes of DN, syndrome of deficiency of Yin and Yang was absent.the proportion of syndrome of deficiency of Qi and Yin was noticeably higher than the other two (P<0.01).In the four accompanied symptoms of DN, blood stasis and damp-heat symptom took the initiative, blood stasis symptom was significantly more than the dampness-heat symptom (P<0.01).
6. In the three deficiency syndromes of DN, Serum hs-CRP increased in syndrome of deficiency of Qi and Yin(3.84±1.93), syndrome of Yin deficiency and dry-heat(2.40±1.06), syndrome of Qi deficiency of spleen and kidney (2.13±0.90) compared with NC (0.86±0.94)(P<0.05, P<0.01). Serum hs-CRP increased in syndrome of deficiency of Qi and Yin compared with syndrome of Yin deficiency and dry-heat, and syndrome of Qi deficiency of spleen and kidney (P<0.05).in the accompanied symptoms of DN, Serum hs-CRP increased in blood stasis (1.83±1.65) and dampness-heat symptom (4.28±2.12) compared with NC (P<0.05, P<0.01), Serum hs-CRP increased in blood stasis symptom compared with dampness-heat symptom (P<0.05)
Conelusion:
1. Serum Fcγ RIIB/C did not change significantly in DN, which could not provide the basis for the diagnosis and treatment of DN;
2. Patients of diabetic nephropathy were in micro-inflammatory state;
3. Serum Fcγ RIIB/C had a weak negative correlation with micro-inflammatory state;
4. Qi and Yin deficiency, accompanied with blood stasis inhibition and dampness-heat was the basic pathogenesis in the early and middle stages of diabetic nephropathy;
5. Serum hs-CRP was related with TCM syndromes, to some extent, which provided an objective basis for the Chinese medicine dialectical type.
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