OBJECTIVE: To establish methods for the identification of Rhizoma Chuanxiong and determination of Ferulic acid( FA) in Ganning granula. METHODS: The identification of Rhizoma Chuanxiong was carried out by TLC and the determination of FA was by performed by HPLC. FA was separated on Kromasil ODS-1 C18 column with water-methanol-acetic acid ( 30∶ 69. 5∶ 0. 5) as mobile phase. The detection wavelength was 322nm, the column temperature was set at room temperature and the flow rate was 1. 0mL? min-1. RESULTS: The TLC spots of the sample presented the same color as its counterparts of Rhizoma Chuanxiong standard and FA reference substance at the corresponding sites. The linear range of FA was 0. 010 12 ～ 0. 151 80? g( r=0. 999 9, n=8) . The average recovery was 97. 39% ( RSD=1. 97% ) . CONCLUSION: The methods of identification and content determination were rapid, accurate, specific and reproducible, and which are suitable for the quality control of Ganning granula.

Objective To investigate the impact of early enteral nutrition on the nutritional status and complications of patients with advanced esophageal carcinomas.Methods Sixty-five patients with advanced esophageal carcinomas were randomly divided into the enteral nutrition group group (n =33) and the control group (n =32).The two groups were given enteral nutrition support and normal nasogastric feeding diet respectively in 24-72 h after hospitalization.The two groups were tested with nutrition indicators:body Mass Index (BMI)/brachial triceps skinfold thickness/upper arm circumference measurement,fasting blood glucose/serum total protein/albumin/cholesterol/triglyceride and the liver function (alanine aminotransferase (ALT)/aspartate aminotransferase(AST)/total bilirubin (TBiL)),and were observed the incidences of complications with liver/intestinal and infection diseases.Results After one month's treatment,compared with the control group,there was significant statistical difference between the two group in patients' nutritional status (BMI index:(22.1 ±4.5) kg/m2 vs.(19.2±4.3) kg/m2; skinfold thickness:(6.2 ±0.4) mm vs.(5.1 ±0.4)mm ; upper arm circumference:(22.8 ± 3.0) cm vs.(20.4 ± 3.2) cm ; serum total protein:(49.2 ± 10.1) g/L vs.(45.1 ± 9.9) g/L; Albumin:(35.5 ± 5.8) g/L vs.(30.6 ± 6.1) g/L; Cholesterol:(5.0 ± 0.6) mmol/L)vs.(4.3 ± 0.7) mmol/L)),the liver function(ALT:(36.0 ± 4.7) U/L vs.(61.5 ± 9.9) U/L; AST:(29.6 ±6.7) U/Lvs.(88.9±10.6) U/L;TBiL:(17.7±3.8) μmol/Lvs.(31.6 ±9.4) μmol/L) (t=2.624,2.036,2.220,2.256,4.155,2.207,2.349,2.476,2.280 respectively,P ＜ 0.05 for all),and the incidence of diarrhea (12％ (4/33) vs.34％ (11/32)) and infection (15％ (5/33) vs.41％ (13/32)) (x2 =2.501,2.193 respectively;P ＜0.05).No statistical difference was observed between the two groups on the levels of serum glucose and triglycerides,and the incidences of complications like bloating/constipation/reflux (P ＞ 0.05).Conclusion The early enteral nutrition could improve the nutritional status of patients with advanced esophageal carcinomas effectively,and reduce the incidence of liver injury,diarrhea and infection.

Oral hydroxycamptothecin nanosuspension (HCPT-Nano) with high supersaturated dissolution level, high permeation and well physical stability, was manufactured by microprecipitation-high press homogenization method. Its pharmaceutical properties were investigated, such as size and distribution, zeta potential, particle shape, physical existence condition, supersaturated dissolution level and so on. Particle size was measured by laser diffraction, and the mean diameters before and after lyophilization were 138 +/- 11.72 nm and 175 +/- 12.74 nm, respectively, for HCPT-Nano. Zeta potentials of HCPT-Nano was over -20 mV. The nanoparticles, being observed by transmission electron microscopy (TEM), were claviform or column in shape. DSC and X-ray diffraction revealed that HCPT existed in the form of crystal for HCPT-Nano. And HCPT-Nano could maintain higher supersaturated dissolution level for long time. So it supplied the possibility of improving oral bioavailability of HCPT when combining together admoveatur of P-gp inhibitor, CsA.

Objective To detect the expression of BRAF V600E mutant protein in cutaneous malignant melanoma (CMM),and to evaluate the sensitivity and specificity of immunohistochemistry (IHC) in detecting BRAF V600E mutation.Methods IHC with an anti-BRAF V600E monoclonal antibody was performed to detect the expression of BRAF V600E mutant protein in paraffin-embedded tissue sections from 103 patients with CMM and 40 patients with nevus.Statistical analysis was carried out with SPSS software version 17.0,and the expression rate of BRAF V600E mutant protein was compared by chi-square test.Results The expression rate of BRAF V600E mutant protein in the CMM patients was 20.4％ (21/103),significantly higher than that in the nevus patients (5.0％ (2/40),x2 =5.06,P ＜ 0.05).Significant differences were observed in the expression rate of BRAF V600E mutant protein between CMM patients of different age groups (29.8％ (14/47) in patients aged ＜ 60 years vs.12.5％ (7/56) in those aged ≥ 60 years,P ＜ 0.05) and nationality (30.2％ (13/43) for Uygur nationality vs.13.3％ (8/60) for Han nationality,P ＜ 0.05),as well as among CMM lesions from different anatomical sites (13.6％ (6/42) in acral sites vs.11.8％ (4/29) in mucous membrane vs.45.8％ (11/32) in non-acral sites,P ＜ 0.05) and of different Clark levels (8.6％ (4/42) for grade Ⅰ-Ⅲ vs.12.4％ (17/61) for grade Ⅳ-Ⅴ,P＜ 0.05),but not between male and female CMM patients or between CMM patients with lymph node metastasis and those without (both P ＞ 0.05).IHC with the anti-BRAF V600E antibody showed a sensitivity of 100％ (15/15) and a specificity of 98.5％ (65/66) in detecting BRAF V600E mutation.Conclusions The expression of BRAF V600E mutant protein is up-regulated in CMM lesions,and CMM patients of Uygur nationality seems to have a higher expression rate than those of Han nationality.IHC appears to be an accurate and rapid method to detect V600E BRAF mutation.

Objective To assess the relationship between BRAF gene mutations and clinical phenotype of malignant melanoma.Methods Tissue specimens were collected from the lesions of 80 patients with malignant melanoma,and from the normal skin of 30 patients with trauma in the Department of Plastic Surgery or General Surgery,and subjected to paraffin embedding and DNA extraction.PCR was performed to amplify the exon 11 and 15 of BRAF gene followed by DNA sequencing.Chi-square test and Fisher's exact test were carried out to assess the relationship between BRAF gene mutations and clinical phenotypes of malignant melanoma.Results BRAF gene mutations were found in 19 (23.8％) of the 80 malignant melanoma specimens.Among the 19 mutationpositive specimens,17 (88.2％) carried mutations in exon 15 of BRAF gene with V600E as the most frequent (88.2％,15/17) mutation type,and 2 (10.5％) carried mutations in exon 11.No mutation was found in any of the normal skin tissue specimens.The average age at onset was 57.5 years in these patients.The frequency of BRAF gene mutation was significantly higher in patients younger than 60 years than in those older than 60 years (37.1％ vs.13.3％,x2=6.613,P ＜ 0.05).A significant difference was observed in the frequency of BRAF gene mutation among tissue specimens of mueosal,acral and non-aeral malignant melanoma (18.2％ (4/21) vs.14.7％(5/34) vs.41.7％ (10/24),x2=6.167,P ＜ 0.05).There was no significant association between BRAF gene mutation and gender,race or lymph node metastasis (all P ＞ 0.05).Conclusions BRAF gene is a hot spot for mutations in patients with malignant melanoma in Xinjiang Uygur Autonomous Region,with V600E point mutation in exon 15 as the most frequent mutation type.BRAF gene mutations appear to be closely correlated with the age at onset of and lesional sites in,but uncorrelated with gender and race of or lymph node metastasis in,patients with malignant melanoma.

OBJECTIVETo study on pharmacokinetics of hydroxycamptothecine (HCPT) nanosuspensions in rats after oral administration.

METHODThe plasma concentrations of HCPT were determined by HPLC-FD. The analysis was performed on a diamonsil C18 column (4.6 mm x 200 mm, 5 microm) with 0.3% acetic acid-triethylamine buffer (pH 5.0) and methanol (57: 43) as mobile phase. The flow rate was 1.0 mL x min(-1); the excitation wave was set at 363 nm, and emission wave was set at 550 nm; the temperature was 35 degrees C. All data of concentration-time of HCPT were treated with pharmacokinetics program DAS 2.0.

RESULTThe concentration-peak area of this assay had a good linear relation in the range from 1 to 50 microg x L(-1), and the minimum limit of quantitation was 1 microg x L(-1). The inter- and intra-day precisions of HCPT were smaller than 4.3%, and the accuracy were between -5.59% and 5.59%. The recoveries of HCPT in three plasma concentrations including high, medial, low concentration were 98.94%, 95.88% and 102.69%, respectively, which was in line with the request of biopharmaceutical analysis. The plasma concentration time profiles of HCPT fitted in two-compartment models well, and the main pharmacokinetic parameters found for HCPT after oral administration were as follows: Cmax 13.10 microg x L(-1), Tmax 0.75 h, t(1/2alpha) 8.242 h, t(1/2beta) 136.122 h, AUC(0-t) 116.77 microg x h x L(-1), AUC(0-infinity) 161.93 microg x h x L(-1).

CONCLUSIONThe HPLC-FD method was simple, with good specificity, reproducibility, and could be used to investigate the pharmacokinetics and determinate the concentration of hydroxycamptothecin. The nanosuspension in this study could accelerate the oral absorption rate of HCPT, and make improving bioavailability of HCPT possible.

ATP-Binding Cassette, Sub-Family B, Member 1 ; antagonists & inhibitors ; Administration, Oral ; Animals ; Calibration ; Camptothecin ; administration & dosage ; analogs & derivatives ; chemistry ; pharmacokinetics ; pharmacology ; Linear Models ; Male ; Nanostructures ; Rats ; Rats, Wistar ; Suspensions

OBJECTIVETo map and identify the disease gene for the epidermolytic palmoplantar keratoderma (EPPK) in a Uighur family of China.

METHODSBlood samples were collected and genomic DNA was extracted from 48 members of the Xinjiang Uighur family. Six microsatellite repeat sequences on chromosome region 17q12-q21 and 12q13 were selected based on the two known candidate genes KRT9 and KRT1. Two-point linkage analysis and haplotype analysis were performed. Exons and their flanking intronic sequence of the KRT9 gene were amplified by polymerase chain reaction (PCR) and sequenced.

RESULTSData from the marker D17S1787 suggested linkage and yielded a Lod score of 8.65 at theta=0 by using MLINK software. Genotypes and haplotypes were acquired. The disease gene of the EPPK family is located between markers 17/TG/36620115 and D17S846. Chromosome 12q13 region was excluded with the negative Lod score obtained in marker D12S96 (Lod=-infinity at theta=0). No pathogenic mutation was detected in the KRT9 gene.

CONCLUSIONThe disease gene of the EPPK family is located on chromosome region 17q21.2. The keratin 9 gene might not be the disease gene.

China ; Chromosomes, Human, Pair 17 ; genetics ; Female ; Humans ; Keratin-1 ; genetics ; Keratin-9 ; genetics ; Keratoderma, Palmoplantar, Epidermolytic ; ethnology ; genetics ; Male ; Microsatellite Repeats ; Mutation ; Pedigree

Objective: To investigate the protective effect and potential mechanism of tanshinol borneol ester (TBE) on homocysteine(Hcy) induced rat bone marrow mesenchymal stem cells (BMSCs) damage. Methods: BMSCs were isolated and cultured in vitro by density gradient centrifugation and adherent culture method. BMSCs were divided into the control (normal isolation and culture), TBE-1(10 μmol/L TBE-1 solution with 100 μl), TBE-2 (10 μmol/L TBE-2 solution with 100 μl), Hcy (0.5 mmol/L Hcy solution with 100 μl), Hcy + TBE-1(0.5 mmol/L Hcy solution with 100 μl, and 10 μmol/L TBE-1 solution with 100 μl), Hcy + TBE-2 (0.5 mmol/L Hcy solution with 100 μl, and 10 μmol/L TBE-2 solution with 100 μl), Hcy+ TBE-1+ inhibitor group(0.5 mmol/L Hcy solution with 100 μl, 10 μmol/L TBE-1 solution with 100 μl, and 25 μmol/L LY294002(specific blocker of phosphatidylinositol 3 kinase) solution with 100 μl), Hcy+ TBE-2+ inhibitor group(0.5 mmol/L Hcy solution with 100 μl, 10 μmol/L TBE-2 solution with 100 μl, and 25 μmol/L LY294002 solution with 100 μl). Cell proliferation activity was detected by MTT assay. The T-SOD activity and malonaldehyde level of cells were measured by anthineoxidase method and TBA method, respectively, to evaluate cell oxidative and antioxidative activities. The ultrastructure of cells was observed under transmission electron microscope. The expression level of PKB and NF-κB of cells in various groups were detected with the immunocytochemical method. Results: (1)Cell proliferation activity in TBE-1 group and TBE-2 group was significantly increased compared with the control group (both P<0.01), and was similar between TBE-1 group and TBE-2 groups after 1, 12, 24 and 48 hours treatment.(2)The T-SOD activity in TBE-1 group and TBE-2 group was significantly higher than in control group (both P<0.01), while it was significantly lower in Hcy group, Hcy+ TBE-1 group, and Hcy+ TBE-2 group than in control group(all P<0.01), and was similar between control group, Hcy+ TBE-1+ inhibitor group, and Hcy+ TBE-2+ inhibitor group(all P>0.05). The T-SOD activity was higher in Hcy+ TBE-1 group and Hcy+ TBE-2 group than in Hcy group(both P<0.01), and was higher in Hcy+ TBE-1+ inhibitor group than in Hcy+ TBE-1 group(P<0.05) and was higher in Hcy+ TBE-2+ inhibitor group than in Hcy+ TBE-2 group(P<0.05). The malonaldehyde level was lower in TBE-1 group and TBE-2 group than in control group(both P<0.01), was higher in Hcy group, Hcy+ TBE-1 group, Hcy+ TBE-2 group, Hcy+ TBE-1+ inhibitor group, and Hcy+ TBE-2+ inhibitor group than in control group(all P<0.01), was lower in Hcy+ TBE-1 group and Hcy+ TBE-2 group than in Hcy group(both P<0.01), was higher in Hcy+ TBE-1+ inhibitor group than in Hcy+ TBE-1 group(P<0.05), was higher in Hcy+ TBE-2+ inhibitor group than in Hcy+ TBE-2 group(P<0.05). (3)Under electron microscope, BMSCs showed profound swelling, senescence and apoptosis of cells increased significantly in Hcy group, Hcy+ TBE-1+ inhibitor group, and Hcy+ TBE-2+ inhibitor group when compared with control group. BMSCs in the TBE-1 and TBE-2 groups presented with abundant rough endoplasmic reticulum, and very active cell metabolism signs. Compared with Hcy group, BMSCs edema, the number of aging and apoptotic cells, and cell injury severity were significantly less in TBE-1+ Hcy group and TBE-2+ Hcy. (4)The PKB level was higher in TBE-1 group and TBE-2 group than in control group(both P<0.01), was lower in Hcy group, Hcy+ TBE-2 group, Hcy+ TBE-1+ inhibitor group, and Hcy+ TBE-2+ inhibitor group than in control group(all P<0.01), was similar between control group and Hcy+ TBE-1 group(P>0.05), was higher in Hcy+ TBE-1 group and Hcy+ TBE-2 group than in Hcy group(both P<0.05), was lower in Hcy+ TBE-1+ inhibitor group than in Hcy+ TBE-1 group(P<0.05), and was lower in Hcy+ TBE-2+ inhibitor group than in Hcy+ TBE-2 group(P<0.05). The NF-κB level was higher in TBE-1 group and TBE-2 group than in control group(both P<0.01), was lower in Hcy group, Hcy+ TBE-1 group, Hcy+ TBE-2 group, Hcy+ TBE-1+ inhibitor group, and Hcy+ TBE-2+ inhibitor group than in control group(P<0.01 or 0.05), was higher in Hcy+ TBE-1 group and Hcy+ TBE-2 group than in Hcy group(both P<0.05), was lower in Hcy+ TBE-1+ inhibitor group than in Hcy+ TBE-1 group(P<0.05) and was lower in Hcy+ TBE-2+ inhibitor group than in Hcy+ TBE-2 group(P<0.05). Conclusion Tanshinol borneol ester can promote the proliferation of BMSC, and attenuate the homocysteine induced rat BMSCs damage possibly through activation of phosphatidylinositol 3 kinase/PKB signal transduction and its downstream signal pathway protein NF-κB.

OBJECTIVE To prepare and characterize curcumin nanomicelles （hereinafter referred to as Cur/mPEG-PBLA micelles）， and to evaluate the in vitro hepatoprotective activity against alcohol liver disease （ALD）. METHODS Cur/mPEG-PBLA micelles were prepared with the dialysis method using methoxy-poly（ethylene glycol）-poly（β-benzyl-L-aspartate） （mPEG-PLGA） as the carrier. The appearance and microscopic morphology of Cur/mPEG-PBLA micelles were observed， and particle size， polydispersity index， Zeta potential， encapsulation efficiency and drug loading content were all detected. The in vitro release， pH stability， thermal stability， dilution stability， storage stability， plasma stability tests， and hemolysis experiments were all performed. The cell model of ALD was established with anhydrous ethanol intervention using human liver cancer cells and normal liver cells as objects， Cur reference solution as reference， to evaluate in vitro preventive and ameliorative effects of Cur/mPEG- PBLA micelles on ALD. RESULTS The prepared Cur/mPEG-PBLA micelles exhibited a pale-yellow milky light， with a spherical shape and uniform distribution. The average particle size was about 140 nm， and the polydispersity index was less than 0.3. Zeta potential was （－8.15±0.05） mV； the encapsulation efficiency was （73.26±3.16）%， and the drug loading content was （4.87± 0.42）%. The cumulative release of Cur reference substance was close to 80% at 10 h； the cumulative release of Cur/mPEG-PBLA micelles at 8 h was 28.94% and only 48.25% at 48 h. pH stability and thermal stability of Cur/mPEG-PBLA micelles were better than those of Cur reference solution； Cur/mPEG-PBLA micelles showed good dilution stability， storage stability and plasma stability， and would not cause hemolysis. Cur reference solution and Cur/mPEG-PBLA micelles had varying degrees of in vitro preventive and ameliorative effects on ALD in two types of cells； after 48 h of application， the above effects of Cur/mPEG-PBLA micelles were significantly better than those of Cur reference solution at the same mass concentration （P＜0.05）. CONCLUSIONS Cur/mPEG-PBLA micelles can improve pH stability and thermal stability of Cur， delayits degradation rate， and have better in vitro hepatoprotective activity against ALD.