[Development of Tianma HPLC fingerprint and discriminant analysis].

Tianma(the tuber of Gastrodia eleta) is a widely used and pricy Chinese herb. Its counterfeits are often found in herbal markets, which are the plant materials with similar macroscopic characteristics of Tianma. Moreover, the prices of Winter Tianma(cultivated Tianma) and Spring Tianma(mostly wild Tianma) have significant difference. However, it is difficult to identify the true or false, good or bad quality of Tianma samples. Thus, a total of 48 Tianma samples with different characteristics(including Winter Tianma, Spring Tianma, slice, powder, etc.) and 9 plant species 10 samples of Tianma counterfeits were collected and analyzed by HPLC-DAD-MS techniques. After optimizing the procedure of sample preparation, chromatographic and mass-spectral conditions, the HPLC chromatograms of all those samples were collected and compared. The similarities and Fisher discriminant analysis were further conducted between the HPLC chromatograms of Tianma and counterfeit, Winter Tianma and Spring Tianma. The results showed the HPLC chromatograms of 48 Tianma samples were similar at the correlation coefficient more than 0.848(n=48). Their mean chromatogram was simulated and used as Tianma HPLC fingerprint. There were 11 common peaks on the HPLC chromatograms of Tianma, in which 6 main peaks were chosen as characteristic peaks and identified as gastrodin, p-hydroxybenzyl alcohol, parishin A, parishin B, parishin C, parishin E, respectively by comparison of the retention time, UV and MS data with those of standard chemical compounds. All the six chemical compounds are bioactive in Tianma. However, the HPLC chromatograms of the 10 counterfeit samples were significantly different from Tianma fingerprint. The correlation coefficients between HPLC fingerprints of Tianma with the HPLC chromatograms of counterfeits were less than 0.042 and the characteristic peaks were not observed on the HPLC chromatograms of these counterfeit samples. It indicated the true or false Tianma can be identified by either the similarity or characteristic peaks on HPLC fingerprint. Comparing the Winter Tianma with Spring Tianma showed that the HPLC chromatograms of 15 winter Tianma samples and 11 spring Tianma samples were similar at the mean correlation coefficient of 0.908. But the intensity of the characteristic peaks were different between the two groups of Tianma samples, i.e. the intensity of gastrodin, paishin A and C in winter Tianma was lower than those in spring Tianma. The Winter Tianma and Spring Tianma could be discriminated by either the Fisher unstandardized discrimination function or Linear discriminant function, based on the peak areas of 11 common peaks on HPLC chromatograms as variate.

[Determination of Bioactive Components in Paeonia lactiflora Roots Cultivated in Various Areas by UHPLC].

Objective: To assess the quality of Paeonia lactiflora roots by the multi-bioactive chemical markers. Methods: Total of66 Paeonia lactiflora roots samples were collected from Sichuan,Zhejiang and Anhui. An UHPLC-DAD technique was employed to quantify the contents of paeoniflorin,hydroypaeoniflorin,abiflorin,pentagalloyglucose,benzoypaeoni-florin,paeonol,gallic and catechin in these samples. These chemical components in each sample were also calculated by Principal Component Analysis( PCA). Results: The eight bioactive components were good separated in 30 min on the UHPLC chromatogram. The correlation coefficients between peak areas and concentration for these bioactive components were not less than 0. 9990( n = 6). And their recoveries were in the range of95. 94% ~ 100. 92%( n = 6). The contents of paeoniflorin in Paeonia lactiflora roots samples collected from Sichuan,Zhejiang and Anhui were 40. 54 mg / g( n = 23),33. 09 mg / g( n = 22) and 39. 47 mg / g( n = 21),respectively. The values of PCA were 0. 4435( n = 23)for the samples from Sichuan,0. 0122( n = 22) for the samples from Zhejiang and- 4. 9850( n = 21) for the samples from Anhui. The content of paeoniflorin in biennial,triennial,four-year,five-year and six-year old Paeonia lactiflora roots were 24. 76( n = 2),37. 17( n= 16),37. 83( n = 23),39. 71( n = 16) and 37. 45 mg / g( n = 7),respectively. Conclusion: The developed method can accurately quantify the content of principal bioactive compounds in Paeonia lactiflora roots. The quality is various among Paeonia lactiflora roots cultivated in Sichuan, Zhejiang and Anhui on the basis of paeoniflorin content or the value of PCA,but the quality of Paeonia lactiflora roots cultivated in Sichuan is the best. Moreover, it is suggested Paeonia lactiflora roots should be harvested in third or fourth year based on the output and quality.

CD40 expression and its prognostic significance in human gastric carcinoma.

This study aimed to detect the relationship between CD40 (protein and mRNA) expression and human gastric cancer and to determine the prognostic significance of CD40 in gastric cancer patients. We collected 128 cases of gastric cancer specimens, and the expression of CD40 (protein and mRNA) was measured by immunohistochemistry and in situ hybridization. Our study indicated that CD40 is constitutively expressed in human gastric carcinoma tissues. Positive expression of CD40 (protein and mRNA) in gastric cancer tissues was closely related to the tumor TNM stage and the presence of distant metastasis, with CD40 mRNA also being correlated with the presence of lymphatic metastasis. Furthermore, the expression of CD40 (protein and mRNA) is closely related to the prognosis of gastric cancer patients. The expression of CD40 protein and mRNA is positively correlated with the presence of distant (for both protein and mRNA) and lymphatic (for mRNA only) metastasis, and an increased tumor TNM stage in gastric carcinoma. Patients who express low levels of CD40 may have a better prognosis than those who have higher levels of CD40.

Transplanted Neural Stem Cells: Playing a Neuroprotective Role by Ceruloplasmin in the Substantia Nigra of PD Model Rats?

Although mounting evidence suggests that ceruloplasmin (CP) deficiency and iron deposition are pivotal factors responsible for exacerbating demise of dopaminergic neurons in the substantia nigra (SN) of the Parkinsonism and neural stem cells (NSCs) are believed to be excellent candidates for compensating the lost dopaminergic neurons, there are few researches to explore the change of CP expression and of iron deposition in the pathological microenvironment of SN after NSCs transplantation and the ability of grafted NSCs to differentiate directionally into dopaminergic neurons under the changed homeostasis. With substantia nigral stereotaxic technique and NSCs transplantation, we found that tyrosine hydroxylase and CP expression decreased and iron deposition increased in the lesioned SN after 6-OHDA administration compared with control, while tyrosine hydroxylase and CP expression increased and iron deposition decreased after NSCs transplantation compared to 6-OHDA administration alone. Only a small number of embedding NSCs are able to differentiate into dopaminergic neurons. These results suggest that grafted NSCs have an influence on improving the content of CP expression, which may play a neuroprotective role by decreasing iron deposition and ameliorating damage of dopaminergic neurons and possibly underline the iron-related common mechanism of Parkinson's disease and Wilson's disease.

Currently Clinical Views on Genetics of Wilson's Disease.

OBJECTIVE: The objective of this study was to review the research on clinical genetics of Wilson's disease (WD). DATA SOURCES: We searched documents from PubMed and Wanfang databases both in English and Chinese up to 2014 using the keywords WD in combination with genetic, ATP7B gene, gene mutation, genotype, phenotype. STUDY SELECTION: Publications about the ATP7B gene and protein function associated with clinical features were selected. RESULTS: Wilson's disease, also named hepatolenticular degeneration, is an autosomal recessive genetic disorder characterized by abnormal copper metabolism caused by mutations to the copper-transporting gene ATP7B. Decreased biliary copper excretion and reduced incorporation of copper into apoceruloplasmin caused by defunctionalization of ATP7B protein lead to accumulation of copper in many tissues and organs, including liver, brain, and cornea, finally resulting in liver disease and extrapyramidal symptoms. It is the most common genetic neurological disorder in the onset of adolescents, second to muscular dystrophy in China. Early diagnosis and medical therapy are of great significance for improving the prognosis of WD patients. However, diagnosis of this disease is usually difficult because of its complicated phenotypes. In the last 10 years, an increasing number of clinical studies have used molecular genetics techniques. Improved diagnosis and prediction of the progression of this disease at the molecular level will aid in the development of more individualized and effective interventions, which is a key to transition from molecular genetic research to the clinical study. CONCLUSIONS: Clinical genetics studies are necessary to understand the mechanism underlying WD at the molecular level from the genotype to the phenotype. Clinical genetics research benefits newly emerging medical treatments including stem cell transplantation and gene therapy for WD patients.

Effect and prognostic significance of the KAI1 gene in human gastric carcinoma.

The present study aimed to explore the effect and mechanism of the Kangai 1 (KAI1) gene in regulating the migration and invasion of gastric carcinoma cells, and the prognostic significance of this gene in gastric cancer patients. Immunohistochemistry and in situ hybridization were used to investigate the role of KAI1 in the progression and prognosis of gastric cancer. The pEGFP-N1-KAI1 plasmid was transfected into human gastric carcinoma SGC7901 cells using liposomes. The effect of transfection with the KAI1 gene was measured using a reverse transcription-semi-quantitative polymerase chain reaction (RT-sqPCR) assay. The Transwell chamber assay was used to study the metastatic and invasive ability of SGC7901 cells. Gastric cancer metastasis-associated genes, including hypoxia-inducible factor (HIF)-1α, matrix metalloproteinase (MMP)-2, MMP-9, basic fibroblast growth factor (bFGF), and urease plasminogen activator (uPA) were measured by RT-sqPCR prior to and following transfection with the KAI1 gene. The expression of KAI1 protein and mRNA was associated with the differentiation degree of gastric cancer, presence of lymph node metastasis, tumor-node-metastasis stage, depth of invasion and the survival time of patients. The migratory and invasive abilities of SGC7901 cells were significantly decreased subsequent to transfection with the KAI1 gene, and the expression of bFGF and uPA was downregulated. It was concluded that the tumor suppressor gene KAI1 inhibits the migration and invasion of gastric carcinoma cells, possibly by suppressing the expression of uPA. Patients that expressed KAI1 may demonstrate an improved prognosis.