ObjectiveTo observe the effect of Buyang Huanwutang in treating diabetic peripheral neuropathy （DPN） by inhibiting pyroptosis through AMP-activated protein kinase （AMPK）/UNC-51-like kinase 1 （ULK1） mitophagy pathway. MethodSixty male SPF SD rats （6-7 weeks old） were used in animal experiments and numbered according to their body mass. They were then randomly divided into four groups by computer： normal group， model group， α-lipoic acid group（60 mg·kg^-1）， and Buyang Huanwutang group（15 g·kg^-1）， with 15 rats in each group. The diabetic model was established by injection of streptozocin （STZ）. After successful modeling， the α-lipoic acid group and the Buyang Huanwutang group were given corresponding drugs， and the normal group and the model group were given normal saline. Sensory nerve conduction velocity （SNCV） and paw withdrawal threshold （PWT） were measured at the end of administration for 12 weeks. Immunohistochemistry and Western blot were used to detect the expression of phosphorylated AMP activated protein kinase （p-AMPK）， phosphorylated UNC-51-like kinase 1 （p-ULK1）， protein involved in microtubule-associated protein 1 light chain 3 （LC3）， selective autophagy receptors （p62/SQSTM1）， Beclin1， NOD receptor protein structure domain-related proteins 3 （NLRP3）， Caspase-1 （Caspase-1）， and interleukin-1 beta （IL-1β） in dorsal root ganglia （DRG）. Immunofluorescence was used to detect the expression of the N-terminal gasdermin D （N-GSDMD）. ResultCompared with those in the normal group， rats in the model group had increased fasting blood glucose （P<0.01） and significantly reduced SNCV， PWT （P<0.01）， LC3Ⅱ/LC3Ⅰ， Beclin1， p-AMPK/AMPK， and p-ULK1/ULK1 （P<0.01）. In addition， p62， NLRP3， N-GSDMD/GSDMD， IL-1β， and cleaved Caspase-1/Caspase-1 were significantly increased （P<0.01）. Compared with those in the model group， SNCV and PWT were increased （P<0.01） in each administration group， and LC3Ⅱ/LC3Ⅰ， Beclin1， p-AMPK/AMPK， and p-ULK1/ULK1 were significantly increased （P<0.05， P<0.01）. p62， N-GSDMD/GSDMD， cleaved Caspase-1/Caspase-1， NLRP3， and IL-1β decreased （P<0.05， P<0.01）. Compared with the α-lipoic acid group， the Buyang Huanwutang group had significantly increased SNCV， PWT （P<0.05）， LC3Ⅱ/LC3Ⅰ， and p-ULK1/ULK1 （P<0.05） and significantly decreased NLRP3 and N-GSDMD/GSDMD （P<0.05）. ConclusionBuyang Huanwutang regulates mitophagy and inhibits pyroptosis through the AMPK/ULK1 pathway to prevent and treat DPN， and its therapeutic effect may be better than α-lipoic acid.

ObjectiveTo investigate the mechanism of Buyang Huanwutang in treating diabetic peripheral neuropathy （DPN） via mitochondrial transport. MethodDiabetes in SD rats was induced by a high-carbohydrate/high-fat diet and intraperitoneal injection of streptozotocin （STZ）. The 45 diabetic rats were randomly assigned into a DPN group， an alpha-lipoic acid （60 mg·kg^-1·d^-1） group， and a Buyang Huanwutang （15 g·kg^-1·d^-1） group， with 15 rats in each group. Fifteen normal SD rats were fed with the standard diet and set as the control group. The rats were administrated with corresponding drugs by gavage for 12 weeks. The paw withdraw threshold （PWT） and motor nerve conduction velocity （MNCV） were measured at the end of medication， and the sciatic nerve and the bilateral dorsal root ganglia of L4-5 were collected. The injury model of NSC34 cells was established by treating with 50 mmol·L^-1 glucose and 250 μmol·L^-1 sodium palmitate. The NSC34 cells were then randomly assigned into a blank （10% blank serum） group， a DPN （10% blank serum） group， an apha-lipoic acid （10% apha-lipoic acid-containing serum） group， a Buyang Huanwutang （10% Buyang Huanwutang-containing serum） group， and a Buyang Huanwutang + Compound C （CC） （10% Buyang Huanwutang-containing serum + 10 μmol·L^-1 CC） group. The cell intervention lasted for 24 h. The immunofluorescence method， immunohistochemistry， and Western blot were employed to determine the expression levels of phosphorylated adenosine monophosphate-activated protein kinase （p-AMPK）， phosphorylated cAMP-response element binding protein （p-CREB）， kinesin family member 5A （KIF5A）， and dynein cytoplasmic 1 intermediate chain 2 （DYNC1I2）. ResultCompared with the control group， the DPN group of rats showed increased fasting blood glucose （P<0.01）， decreased MNCV and PWT （P<0.01）， down-regulated expression of KIF5A， p-AMPK/AMPK， and p-CREB/CREB （P<0.01）， and up-regulated expression of DYNC1I2 （P<0.01）. Compared with the DPN group， drug intervention groups showed increased MNCV and PWT （P<0.01）， up-regulated expression of KIF5A， p-AMPK/AMPK， and p-CREB/CREB （P<0.05， P<0.01）， and down-regulated expression of DYNC1I2 （P<0.05， P<0.01）. The Buyang Huanwutang group had higher levels of MNCV and KIF5A （P<0.05） and lower level of DYNC1I2 （P<0.01） than the apha-lipoic acid group. Compared with the blank group， the DPN group of NSC34 cells showed decreased levels of KIF5A， p-AMPK/AMPK， and p-CREB/CREB （P<0.01） and increased level of DYNC1I2 （P<0.01）. The apha-lipoic acid group and Buyang Huanwutang group had higher levels of KIF5A， p-AMPK/AMPK， and p-CREB/CREB （P<0.05， P<0.01） and lower level of DYNC1I2 （P<0.01） in NSC34 cells than the DPN group. Buyang Huanwutang group had higher KIF5A level （P<0.05） in NSC34 cells than the apha-lipoic acid group. Moreover， the Buyang Huanwutang + CC group had lower levels of KIF5A， DYNC1I2， p-AMPK/AMPK， and p-CREB/CREB （P<0.01） in NSC34 cells than the Buyang Huanwutang group. ConclusionBuyang Huanwutang may regulate mitochondrial anterograde transport via the AMPK/CREB pathway to prevent and treat DPN.

Objective:To explore effects of T helper type 1 cells (Th1) to T helper type 2 cells (Th2) ratio and the related cytokines on the prognosis of patients with colorectal neoplasms.Methods:A total of 98 colorectal neoplasms patients undergoing the surgery admitted in Suqian Hospital Affiliated to Xuzhou Medical University from December 2015 to December 2017 were enrolled, and all patients were selected as the colorectal cancer group. According to Dukes staging criteria, patients were divided into stage A (25 cases), stage B (30 cases) and stage C (43 cases). In addition, 72 healthy subjects who underwent physical examination in Suqian Hospital Affiliated to Xuzhou Medical University during the same period were selected as the healthy control group. Preoperative venous blood on an empty stomach was extracted from the healthy control group and the colorectal cancer group. Flow cytometry was used to analyze the Th1/Th2 ratio in peripheral blood. The levels of cytokines interferon (IFN)-γ, interleukin (IL)-2, IL-4 and IL-10 in serum samples were detected by using enzyme-linked immunosorbent assay (ELISA). After operation, patients were followed up by telephone or outpatient service. The Th1/Th2 ratio and levels of IFN-γ, IL-2, IL-4 and IL-10 of both groups at different stages of both groups were compared. The correlation between Th1/Th2 ratio and the clinicopathological characteristics of colorectal cancer patients was analyzed. Kaplan-Meier method was used to make survival analysis and Cox regression model was used to analyze influencing factors for overall survival (OS).Results:The Thl/Th2 ratio in colorectal cancer patients was lower than that in the healthy control group (5.13±2.04 vs. 11.82±2.76, t = 18.177, P < 0.01). The lymphovascular invasion and Dukes stage C ratio in patients with decreased Th1/Th2 ratio were higher than those in patients with increased Th1/Th2 ratio ( χ2 values were 16.403, 16.248, both P < 0.01). The levels of IFN-γ and IL-2 in serums of colorectal patients were (95±15) ng/L and (78±10) ng/L, respectively, which were lower than those in the healthy control group [(157±17) ng/L and (123±12) ng/L, t values were 25.160, 26.622, all P < 0.01]. The levels of IL-4 and IL-10 in the colorectal cancer group were (87±16) ng/L and (178±18) ng/L, respectively, which were higher than those in the healthy control group [(46±9) ng/L and (124±12) ng/L] ( t values were 19.577, 22.095, all P < 0.01). The follow-up time ranged from 31.0 to 55.0 months, and the median follow-up time was 37.2 months and the median OS time was 21.0 months. Survival analysis showed that the OS of patients with increased Th1/Th2 ratio was better than that of patients with reduced Th1/Th2 ratio ( χ2 = 7.287, P = 0.007). Multivariate Cox regression analysis showed that lymph node metastasis, tumor stage, and Th1/Th2 ratio were independent influencing factors for OS in colorectal cancer patients ( OR values were 8.541, 3.442, 1.275, all P < 0.05). Conclusion:The imbalance of related cytokines secreted by Th1 and Th2 cells and the decrease in the ratio of Th1/Th2 are related to the progression and the poor prognosis of colorectal cancer.

Objective To analyze the angiographic results of intracranial aneurysms without complete embolization immediately after low-profile visualized intraluminal support device (LVIS) stent at mid-term follow up,and explore the risk factors for healing of partial occluded aneurysms.Methods One hundred and sixty-one patients with intracranial aneurysms treated by LVIS stent embolization in our hospital from December 2014 to December 2018 were selected;193 aneurysms in total,including 93 un-ruptured aneurysms and 100 ruptured aneurysms,were noted.DSA was performed immediately after operation to evaluate the degrees of aneurysm embolization according to Raymond grading criteria.The embolization degrees of aneurysms were compared at 8 months after surgery and immediately after surgery,and the healing of aneurysms (Raymond grading 1) was calculated.Univariate Logistic regression analysis and multivariate Logistic regression analysis (forward maximum likelihood ratio method) were used to screen the risk factors for healing of incomplete aneurysm embolization.Results The embolization degrees immediately after surgery were as follows:78 were with Raymond grading 1,54 with Raymond grading 2,and 61 with Raymond grading 3;complete aneurysm embolization were noted in 78 aneurysms (40.41％) and incomplete aneurysm embolization were noted in 115 (59.59％).Follow up (8 months after surgery) results indicated that,of the 193 aneurysms,171 were with Raymond grading 1,10 with Raymond grading 2,and 12 with Raymond grading 3;there were 171 aneurysms (88.60％) having complete aneurysm embolization and 22 (11.40％) having incomplete aneurysm embolization.In the aneurysms having incomplete aneurysm embolization immediately after surgery,the healing rate was 81.74％ (94/115).Univariate Logistic regression analysis showed that hypertension,diabetes mellitus,posterior circulation aneurysm,dissecting aneurysm,body-neck ratio and embolization degrees immediately after surgery were risk factors for healing of aneurysms (P＜0.05).Multivariate Logistic regression analysis revealed that posterior circulation aneurysm and diabetes mellitus were independent risk factors for healing of aneurysms (P＜0.05).The area under the curve of receiver operating characteristic curve of the regression model was 0.755,indicating that the prediction efficiency of the regression model was moderate.Conclusion Treatment of intracranial aneurysms with LVIS stent is effective,and the complete occlusion rate is high,even in the aneurysms with incomplete embolization immediately after surgery;aneurysms located in the posterior circulation and associated with diabetes can affect the treatment of aneurysms.

OBJECTIVE:To develop and improve the quality standard for Kanggu zengsheng tablet. METHODS:TLC was used for the qualitative identification of Drynaria fortunei,Epimedii Folium and Spatholobus suberectus;HPLC was used for the contents determination of icariin and acteoside:the column was Diamonsil C18 with mobile phase of acetonitrile-0.1% formic acid solution (gradient elution) at a flow rate of 1.0 mL/min;detection wavelength was 270 nm for icariin and 334 nm for acteoside, Cdumn temperature was 25 ℃,and the injection volume was 10 μL. RESULTS:The TLC spots of D. fortunei,Epimedii Folium and S. suberectus were clear and well separated,negative control without interference. The linear range was 0.0188-1.88 μg for ac-teoside(r=0.9999)and 0.107-2.14 μg for icariin(r=0.9999);RSDs of precision,stability and reproducibility tests were lower than 2.0%;recoveries were 100.2%-105.0%(RSD=1.6%,n=9) and 96.2%-99.5%(RSD=1.4%,n=9). CONCLUSIONS:The improved standard can more effectively control the quality of Kanggu zengsheng tablet.

Objective To predict the disease progression risks of healthy rhesus ( normal) and rhesus infected with simian immunodeficiency virus ( SIV) in the stages of long-term nonprogressor ( LTNP) , normal progressor ( NP) , rapid progressor ( RP) by discriminant analysis. Methods Five-year observation was carried out in SIV infected rhesus model without any intervention. The SIV infected rhesus model at the stages of LTNP, NP, RP were selected, 10 in each group, and T lymphocyte subsets and serum parameters for spleen-deficiency syndrome and kidney-deficiency syndrome in SIV infected rhesus were compared with 5 healthy monkey having the same survival time. The influence factors of different types of disease progression were screened from T cell subsets and Chinese medical syndrome indexes, and then the discriminant equation was established to predict the risks. Results White blood cell ( WBC) count and lymphocyte ( LYM) ratio were enrolled into the discriminant equation before infection, and T4 level and Log10RNA of set point were enrolled into the discriminant equation in the platform period. The test results for the uniform rate of the established discriminant function showed that the total coincidence rate of theoretic distinguish to the actual data was 57.1% , 91.2%respectively before infection and in the platform period. Conclusion The pre-infection WBC count and LYM ratio can be used as a reference for the evaluation of different types of disease progresson, and Log10RNA and T4 level at platform phase can be used as the predicting factors of different types of disease progression risk prediction.

DNA barcoding technology is widely opplied for species identification and discovery by using short and standard fragments of DNA sequences. In this study, the cytochrome c oxidase subunit 1 (COI) sequence as a DNA barcode was used to identify the Gekkogecko Linnaeus medicinal materials and adulterants in order to provide a new identification method of G. gecko Linnaeus. Genomic DNA was extracted from the experimental samples. The COI regions of nrDNA were amplified using polymerase chain reaction and sequenced bidirectionally. The alignment and NJ tree construction were performed in MEGA6.0. COI sequences can be obtained from all samples. The average intra-specific K2P distance of G. gecko Linnaeus was 0.005 and the maximum intra-specific distance was 0.013. All the G. gecko Linnaeus medicinal samples clustered into a clade in the NJ tree and can be distinguished from the adulterants. In a conclusion, COI can be used to correctly identify G. gecko Linnaeus, and it will be a potential DNA barcode for identification of other animal medicinal materials.

Objective: This study aimed to distinguish Serpentis Periostracum from its adulterants, which will provide the basis for its safe application. Methods: Here, COI sequences of 68 samples from 13 species were PCR amplified and sequenced. Furthermore, the DNA Barcoding Gap and phylogenetic cluster analysis were carried out. Results:The results exhibited that the COI sequences of all the three origin animals of Serpentis Periostracum have DNA Barcoding Gap. For phylogenetic cluster analysis, all the three origin animals showed monophyletic and every species can be discriminated clearly. Conclusion: COI is an effective DNA barcode for the identification of Serpen-tis Periostracum.

Objective To detect the changes of peripheral blood T lymphocyte subsets in patients with hand,foot and mouth disease (HFMD) and investigate its clinical significance.Methods Three hundred and eighty-six patients with HFMD and 78 healthy children were detected the peripheral blood T lymphocyte subsets levels using the flow cytometry (FCM).The patients were divided into 325 common patients and 61 critically ill patients according to the severity of disease.Among the 386 patients,264 were in infection stage and 122 were in recovery stage.Compared the differences of peripheral blood T lymphocyte subsets levels in patients and healthy controls,common patients and critically ill patients,infection patients and recovery patients respectively.Results The patients' numbers of CD3 +,CD4 +,CD8 + cells and CD4 +/CD8 + ratio were (47.4 ± 10.6) ％,(24.0 ±8.0) ％,(23.2 ± 7.4) ％ and 1.0 ± 0.5 respectively.These parameters were significantly different with those in the control group ((68.4±7.5)％,(38.2 ±6.9)％,(18.8 ±4.6)％ and 2.0 ±0.4 respectively;t =3.16,4.62,2.23 and 2.51 respectively,P ＜ 0.05).The common patients' numbers of CD3 +,CD4 +,CD8 + cells and CD4+/CD8 + ratio were (53.0 ±9.8)％,(26.5 ±7.9)％,(20.4±7.5)％and 1.3 ±0.6,which had significant differences compared with those of the critically ill patients ((42.3 ± 10.9) ％,(18.7 ± 4.7) ％,(24.2 ± 7.4) ％ and (0.7 ± 0.4) respectively; t =2.25,2.47,1.91 and 2.32 respectively,P ＜ 0.05).The infection patients' CD3 +,CD4 +,cell numbers and CD4 +/CD8 + ratio were (45.3 ± 10.5) ％,(21.6 ± 8.4) ％and 0.9 ± 0.5.These parameters were significantly different with those of the recovery patients ((54.7 ±9.9)％,(27.5 ± 9.1)％ and 1.4 ± 0.6 respectively; t =2.41,1.87 and 2.92 respectively,P ＜ 0.05).Conclusion The cellular immunity of patients with HFMD was impaired.The T lymphocyte function was suppressed,and the level of suppression was positively correlated to severity of the illness.

Medicinal animals are important part of Traditional Chinese medicine resources in China. Cytochrome c oxidase subunit I (COI) was selected as the standard DNA barcoding sequence for animal medical materials. In this study, the 51 animal species from 45 animal medical materials in the Chinese Pharmacopoeia were selected and the intra-specific variation and the inter-specific divergence, the barcoding gap, the identification efficiency of their COI sequences were analyzed. The results showed that the inter-specific divergence is higher than intra-specific distance. The barcoding gap existed between inter-specific sequence divergence and intra-specific dis-tance. The identification efficiencies were 100% both at the genus and species level except the Arthropoda. The cluster dendrogram exhibited that different species distinguished from others. Therefore, COI sequence as a bar-code is suitable to identify the species of animal medical materials in Chinese Pharmacopoeia.