OBJECTIVES: To investigate the effect of Didang Decoction-medicated serum on autophagy in high glucose (HG)-induced rat glomerular endothelial cells (RGECs) and explore the pathway that mediates its effect. METHODS: Primary RGECs were isolated and cultured using sequential sieving combined with collagenase digestion, followed by identification using immunofluorescence assay for factor VIII. High glucose medium was used to induce RGECs to simulate a diabetic environment, and the effects of Didang Decoction-medicated serum and 3-MA (an autophagy inhibitor), either alone or in combination, on autophagy of HG-exposed cells were evaluated by observing autophagic vacuoles using monodansylcadaverine (MDC) staining. RT-qPCR and Western blotting were employed to measure mRNA and protein expression levels of Beclin-1, p62, LC3B, p-PI3K, p-Akt, and p-mTOR. RESULTS: Compared with the control cells, the HG-exposed RGECs showed significantly reduced autophagic fluorescence intensity, decreased Beclin-1 mRNA expression, increased p62 mRNA expression, downregulated Beclin-1 protein and LC3-II/I ratio, and upregulated p62, p-PI3K, p-Akt, and p-mTOR protein levels. Didang Decoction-medicated serum significantly enhanced autophagic fluorescence intensity in HG-exposed cells, increased Beclin-1 mRNA expression, decreased p62 mRNA expression, upregulated Beclin-1 protein, and downregulated p62, p-PI3K, p-Akt, and p-mTOR protein levels. CONCLUSIONS Didang Decoction-medicated serum enhances autophagy in HG-exposed RGECs by regulating the PI3K/Akt/mTOR signaling pathway, which sheds light on a new therapeutic strategy for diabetic nephropathy.
Animals ; Autophagy/drug effects* ; Signal Transduction/drug effects* ; Rats ; TOR Serine-Threonine Kinases/metabolism* ; Drugs, Chinese Herbal/pharmacology* ; Proto-Oncogene Proteins c-akt/metabolism* ; Endothelial Cells/metabolism* ; Phosphatidylinositol 3-Kinases/metabolism* ; Glucose ; Cells, Cultured ; Kidney Glomerulus/cytology* ; Rats, Sprague-Dawley

This study investigates the clinical differentiation and treatment strategies for bipolar disorder （BD） by analyzing the relationship of its core pathomechanisms including qi stagnation， blood stasis， phlegm turbidity， and heat constraint with bile acid metabolism. The imbalance of "yin in form and yang in function" leads to qi stagnation， blood stasis， phlegm turbidity， and heat constraint， which are critical in the pathogenesis and progression of BD. Bile acids regulate neuroinflammation， neural plasticity， and intestinal flora homeostasis through receptor-mediated pathways. It is believed that the physiological functions of bile acids concretely embody the concept of the "liver is yin in form and yang in function" theory. Clinically， prescriptions such as Sini Powder （四逆散） with the function of venting pathogen and resolving constraint， Wendan Decoction （温胆汤） of drying dampness and resolving phlegm， Longdan Xiegan Decoction （龙胆泻肝汤） of clearing liver and draining fire， and Huanglian Ejiao Decoction （黄连阿胶汤） of nourishing yin and blood can be used to nourish liver yin and restore liver yang function. These strategies may improve BD prognosis by modulating bile acid synthesis and metabolism.

To summarize XU Jingshi's clinical experience in treating fever from the perspective of latent pathogen. It is believed that latent pathogen heat accumulation is an important pathogenesis of fever. For latent pathogen in shaoyang and membrane-source， modified Xiaochaihu Decoction （小柴胡汤） plus Qinghao （Artemisiae Annuae Herba） could be used to expel pathogen through membrane and regulate shaoyang； for latent pathogen leading to yin fluids consumption， treatment should clear heat and expel pathogen， and prescribed with Chaihu （Bupleuri Radix）， Huangqin （Scutellaria baicalensis Georgi）， Qinghao （Artemisia carvifolia） combined with Baihu Decoction （白虎汤） or Zhuye Shigao Decoction （竹叶石膏汤） or Qinghao Biejia Powder （青蒿鳖甲散） or Shengmai Decoction （生脉饮） or Siyin Decoction （四阴煎） by syndrome differentiation； for latent pathogen leading to liver wind or reverse transmission to the pericardium， treatment should cool to open pathways， and prescribed with modified Angong Niuhuang Pill （安宫牛黄丸） to clear mind and open the orifices， clear heat and resolve toxins， resolve constraint with aromatics， and expel pathogens to outside； for fever after radiotherapy， chemotherapy or surgery for tumours， prescribed with self-prescribed Fuzheng Anzhong Decoction （扶正安中汤） plus Xiaochaihu Decoction （小柴胡汤） and modified Qinghao Biejia Decoction （青蒿鳖甲汤） to reinforce healthy qi and dispel pathogen， and support the middle jiao.

ObjectiveTo observe the protective effect of Didang Xianxiong decoction on the kidneys of diabetic rats， its regulation on the phosphatidylinositol 3-kinase （PI3K）/protein kinase B （Akt） signaling pathway， and its influence on podocyte apoptosis and explore the mechanism of Didang Xianxiong decoction in improving diabetic nephropathy. MethodThe diabetic model was established by a single intraperitoneal injection of streptozotocin （STZ） solution of 55 mg·kg^-1. The successfully replicated model rats were randomly divided into the model group， Didang Xianxiong decoction group （8.10 g·kg^-1）， Xiao Xianxiongtang group （4.05 g·kg^-1）， Didangtang group （4.05 g·kg^-1）， and alagebrium （ALT-711） group （3 mg·kg^-1）， with six rats in each group. In addition， six rats were included in the blank group. After continuous administration for eight weeks， hematoxylin-eosin （HE） staining was used to observe the pathological changes in rats' kidney tissue. Masson staining was used to observe the degree of collagen deposition. Periodic acid-Schiff （PAS） staining was used to observe basement membrane lesions， and immunohistochemistry was used to detect the expression of phosphorylation （p）-PI3K and p-Akt proteins in rats' kidney tissue. The terminal deoxynucleotidyl transferase-mediated dUTP nick-end labeling （TUNEL） method was used to detect podocyte apoptosis. Real-time fluorescence quantitative polymerase chain reaction （Real-time PCR） was used to detect the mRNA expression of PI3K and Akt in rats' kidney tissue. Western blot was used to detect the protein expression of PI3K， p-PI3K， Akt， p-Akt， B-cell lymphoma 2 （Bcl-2）， Bcl-2-associated X protein （Bax）， phosphorylation glycogen synthase kinase-3β （p-GSK-3β）， and Caspase-3 in the kidney tissue. ResultCompared with the normal group， the model group had compensatory expansion of glomeruli， proliferation of mesangial cells， a large amount of collagen deposition in the mesangial stroma， thickening of the basement membrane， decreased mRNA expression of PI3K and Akt， and inhibition of PI3K and Akt protein phosphorylation （P<0.01）. It also underwent enhanced apoptotic signaling， decreased expression of anti-apoptotic protein Bcl-2 （P<0.01）， and increased expression of Bax， p-GSK-3β， and Caspase-3 （P<0.01）. Compared with the model group， Didang Xianxiong decoction significantly improved kidney tissue pathology， increased mRNA expression of PI3K and Akt （P<0.01）， significantly up-regulated phosphorylation levels of PI3K and Akt proteins （P<0.01） and Bcl-2 expression （P<0.01）， downregulated the expression of Bax， p-GSK-3β， and Caspase-3 （P<0.01）， and weakened podocyte apoptotic signaling. ConclusionDidang Xianxiong decoction may promote the activation of the PI3K/Akt signaling pathway， inhibit podocyte apoptosis， and thus slow down the progression of diabetic nephropathy.

Objective:To observe the intervention mechanism of phlegm-stasis co-treatment for the JNK signaling pathway in the myocardium of diabetes rats.Methods:Totally 50 male SD rats of SPF grade were selected. Diabetes model was established by single intraperitoneal injection of 55 mg/kg streptozotocin (STZ) solution. After continued feeding for 3 weeks, the rats were divided into normal group, model group, alachloramine group, blood stasis removing group, phlegm removing group and phlegm-blood stasis co-treatment group according to random number table method, with 6 rats in each group. Xiaoxianxiong Decoction (4.05 g/kg), Xuefu Zhuyu Decoction (7.02 g/kg), Didang Xianxiong Decoction (8.10 g/kg) were administered to the stomach respectively in the phlegm removing group, the blood stasis removing group and the phlegm-blood stasis co-treatment group. Alachloramine (3 mg/kg) was administered to the stomach by gavage in the alachloramine group. After 8 weeks, HE staining was used to observe the morphological changes of myocardial tissue in diabetic rats. Masson staining was used to observe the deposition of collagen fibers in the myocardial interstitium in rats. The expression of JNK1 protein was determined by immunohistochemistry. JNK1 mRNA, IRS1 mRNA and NLRP3 expression levels were detected by Real-time PCR. Western blot was used to detect the protein expressions of IRS-1, p-Akt and NLRP3.Results:The myocardial cells in the model group were disorganized, with hypertrophy, blurred texture, inflammatory infiltration of interstitium, increased collagen fibers, and focal necrosis. All treatment groups could improve fibrosis, inflammatory infiltration and reduce myocardial collagen deposition in different degrees. Compared with the model group, the mRNA and protein expressions of JNK1 and NLRP3 bodies decreased ( P<0.01), the IRS-1 mRNA and protein increased ( P<0.01), and p-Akt protein expression increased ( P<0.01). Conclusions:The phlegm and stasis co-treatment can effectively improve the cardiomyopathy of diabetes rats, and the effect is better than the phlegm-resolving method or the stasis resolving method alone. The mechanism may be related to the inhibition of JNK signaling pathway activation, reduce the expressions of JNK1 and NLRP3, and increase the IRS-1 and Akt.

ObjectiveTo observe the effect of Didang Xianxiong decoction on the ultrastructure of myocardium and the expression of connective tissue growth factor （CTGF） and low-density lipoprotein receptor-related protein （LRP） in diabetic rats. MethodThe diabetic rat model was established by the intraperitoneal injection of high-dose streptozotocin （55 mg·kg^-1） and the rats were further fed for 3 weeks. Forty model rats were randomly assigned into model group， a Xiao Xianxiongtang （4.05 g·kg^-1·d^-1） group， Xuefu Zhuyutang （6.30 g·kg^-1·d^-1） group， Didang Xianxiong decoction （8.10 g·kg^-1·d^-1） group， and alagebrium chloride （ALT-711， 3 mg·kg^-1·d^-1） group， with 8 rats in each group. Ten normal healthy rats were randomly selected as the blank group. The corresponding drugs were administrated by gavage， and the blank group and the model group were given distilled water at a dose of 10 mL·kg^-1·d^-1 for 8 weeks. The myocardial tissue was collected from the rats under anesthesia at the end of the 8^th week for pathological examination. The expression of CTGF and its receptor LRP in the myocardial tissue was detected by immunohistochemistry and Western blot. ResultCompared with the blank group， the modeling led to obvious ultrastructural damage of the myocardium， up-regulated the expression of CTGF （P<0.01）， and did not significantly change the expression of LRP. Compared with the model group， the drugs alleviated the damage in myocardium ultrastructure， down-regulated the expression of CTGF （P<0.01）， and did not significantly change the expression of LRP. Moreover， Didang Xianxiong decoction showed better performance than Xiao Xianxiongtang and Xuefu Zhuyutang （P<0.01）. ConclusionDidang Xianxiong decoction may protect myocardial tissue by down-regulating the high expression of CTGF in myocardial tissue of diabetic rats， thereby delaying myocardial fibrosis. The results indicate that the therapy of treating both phlegm and blood stasis has better performance than the simple method of resolving phlegm or stasis.

OBJECTIVE: To observe the effect of traditional Chinese medicine for intervention of phlegm and blood stasis in regulating TGF-β1/Smad3 signaling and relieving nephropathy in diabetic rats. METHODS: SD rats were divided into blank group (NC), diabetic model group (MC group), intervention of phlegm and blood stasis (RPDBS) group, phlegm-removing (RP) group and blood-removing (DBS) group. Diabetic models were established in all the rats except for those in the blank group. After 4 weeks of feeding, the rats in RPDBS group, RP group and DBS group were given corresponding drug intervention for 8 weeks. HE staining was used to observe the changes in renal histopathology. Western blotting and real-time fluorescence quantitative PCR were used to detect the expression levels of transforming growth factor-β1 (TGF-β1) and Smad3. RESULTS: The structure and arrangement of the glomeruli and renal tubules improved significantly in the treatment groups in comparison with those in the MC group. The expression levels of TGF-β1, Smad3 and p-Smad3 were significantly downregulated at both the protein and mRNA levels in the treatment groups ( < 0.05), and the down-regulation was more obvious in RPDBS group than in RP group and DBS group ( < 0.05). CONCLUSIONS Intervention of phlegm and blood stasis may inhibit the activation of TGF-β1/Smad3 signaling pathway and delay diabetic nephropathy and fibrosis to protect the renal function in diabetic rats.
Animals ; Diabetes Mellitus, Experimental ; Diabetic Nephropathies ; Kidney ; Rats ; Rats, Sprague-Dawley ; Signal Transduction ; Smad3 Protein ; Transforming Growth Factor beta1

OBJECTIVE: To observe the effects of a traditional Chinese medicine (TCM) capsule for replenishing qi, nourishing yin and activating blood on Notch/Hes1 signaling pathway in the renal tissue and vascular endothelial CD34 and CD144 expressions in a rat model of diabetic nephropathy. METHODS: Rat models of early-stage diabetic nephropathy were established by left nephrectomy and high- fat and high- sugar feeding combined with intraperitoneal injection of STZ. The rats were randomized into model group, benazepril group, and high-, moderate-, and low-dose TCM capsule groups for corresponding treatments, with 6 normal rats as the control group. After 8 weeks of drug treatment, blood glucose and 24-h urinary albumin of the rats were measured, and the renal histopathology was observed with HE staining; Hes1 expression in the renal tissue was detected with immunohistochemical staining, and the renal expressions of CD34 and CD144 were detected using Western blotting. RESULTS: Compared with the normal control group, the rat models of diabetic nephropathy showed obvious abnormalities in 24- h urinary albumin and expressions of Hes1, CD34 and CD144d. The TCM capsule at both the high and moderate doses significantly reduced 24-h urinary albumin in the rats; the renal expressions of Hes1 and CD34 was significantly reduced in all the dose groups, and the expression of CD144 was significantly reduced in the high- dose group. Compared with benazepril group, the TCM capsule obviously reduced CD34 expression at all the 3 doses and lowered CD144 expression at the low dose. Histopathologically, the rats in the model group showed glomerular hypertrophy, increased mesenteric matrix, thickening and widening of the mesenteric membrane, and nodular hyperplasia. These pathologies were obviously alleviated by treatment with the TCM capsule at the high and moderate doses. CONCLUSIONS The Traditional Chinese medicine (TCM) capsule for replenishing qi, nourishing yin and activating blood can reduce Hes1, CD34 and CD144 in kidney tissue of model rats, play a protective role on kidney function and delay the development of DN.
Animals ; Diabetic Nephropathies ; Drugs, Chinese Herbal ; Medicine, Chinese Traditional ; Qi ; Rats ; Signal Transduction ; Transcription Factor HES-1

Diamicron group. Conclusion Modified Taohe Chengqi Decoction and its diferent extractions maybe effective on raising or stabilizing the activity of Na+-K+-ATPase and Ca2+-ATPase in cadiocyte of diabetic rats, so it could prevent myocardium hypertrophy and patho-change by heart function failure in variably extent. Ethyl-acetate group and n-butyl alcohol group showed rather good effect among these extractions.