[SNP Panel Analysis of Ancestry Inference in East Asian Populations].

ABSTRACT: Objective To develop an SNP Panel for East Asian population, which has a high individual identification rate and the capability of ancestry analysis. Methods The 55 SNP Panel by Professor KIDD of Yale University and the 128 SNP Panel by Professor SELDIN of Davis School of California University, 170 SNP Panel in total was used as the basis and its test data in the East Asian population was collected. The genetic parameters of SNP loci were calculated and combined with the results of heatmap analysis to screen SNP loci suitable for East Asian population. Some Tibetan and Han samples were tested. The possibility of using the SNP loci in ancestry inference was analyzed by means of STRUCTURE analysis, principal component analysis and heatmap analysis. Results A Panel with 45 SNPs （45 SNP Panel） was screened out, and the average genetic parameters of each SNP were better than 170 SNP Panel, with the same ancestry analysis and inference ability. Conclusion In terms of ancestry inference information, the 45 SNP Panel can completely replace the 170 SNP Panel and achieve the same ancestry analysis and inference ability. In genetic parameters, 45 SNP Panel is better than 170 SNP Panel in the East Asian population, which shows its important potential forensic application value.

Feature extraction method for proteins based on Markov tripeptide by compressive sensing.

BACKGROUND: In order to capture the vital structural information of the original protein, the symbol sequence was transformed into the Markov frequency matrix according to the consecutive three residues throughout the chain. A three-dimensional sparse matrix sized 20 × 20 × 20 was obtained and expanded to one-dimensional vector. Then, an appropriate measurement matrix was selected for the vector to obtain a compressed feature set by random projection. Consequently, the new compressive sensing feature extraction technology was proposed. RESULTS: Several indexes were analyzed on the cell membrane, cytoplasm, and nucleus dataset to detect the discrimination of the features. In comparison with the traditional methods of scale wavelet energy and amino acid components, the experimental results suggested the advantage and accuracy of the features by this new method. CONCLUSIONS: The new features extracted from this model could preserve the maximum information contained in the sequence and reflect the essential properties of the protein. Thus, it is an adequate and potential method in collecting and processing the protein sequence from a large sample size and high dimension.

[Effects of Huanglian ointment on wound healing of mice with full-thickness skin defect and the related mechanism].

Objective: To observe the effects of Huanglian ointment on wound healing of mice with full-thickness skin defect, and to explore the related mechanism. Methods: Thirty male C57BL/6J mice were divided into Huanglian ointment group and vehicle group according to the random number table after round wounds of full-thickness skin defect with diameter of 7.5 mm were inflicted on the back of each mouse, with 15 mice in each group. Wounds of mice in Huanglian ointment group and vehicle group were treated with Huanglian ointment and vehicle respectively from post injury day (PID) 1 on, 2 times each day. Five mice from each group were selected to observe wound changes on PID 0, 3, 7, 10, and 14, and wound healing rates were calculated. Five mice out of the 10 mice that hadn't been used for general observation in each group were sacrificed on PID 3 and 7 respectively, and 5 mice after being used for general observation in each group were sacrificed on PID 14. Wound and skin tissue within 2 mm from the edge of wound was collected. Histologic scoring was conducted based on the histomorphological observation with HE staining. The expression of double positive cells of alpha smooth muscle actin (α-SMA) and Ki-67 (myofibroblast) in tissue of wounds of mice was observed by immunofluorescence staining. Protein expressions of transforming growth factor beta (TGF-ß) and collagen in tissue of wounds of mice were determined by enzyme-linked immunosorbent assay. Data were processed with analysis of variance for repeated measurement, analysis of variance of factorial design, t test of two independent samples, one-way analysis of variance, and Bonferronni test or correction. Results: (1) Wounds of mice in two groups were red and swollen on PID 0, while they were neither red nor swollen with scabs on PID 3 and 7. On PID 10, woundsof mice in Huanglian ointment group contracted obviously, while the contracted wounds of mice in vehicle group were smaller than those in Huanglian ointment group. On PID 14, wounds of most mice in Huanglian ointment group were healed, while wounds of some mice in vehicle group failed to heal. Wound healing rates of mice in two groups were close on PID 3 and 7 (with t values respectively 0.64 and 1.90, P values above 0.05). Wound healing rates of mice in Huanglian ointment group on PID 10 and 14 were (76±7)% and (93±5)% respectively, significantly higher than those of vehicle group [(48±9)% and (68±11)%, with t values respectively 7.44 and 3.89, P values below 0.01]. Wound healing rates of mice in two groups on PID 7, 10, and 14 were significantly higher than those on the previous time points of the same group (with P values below 0.01). (2) Histologic scores of wounds of mice in two groups were close on PID 3 (t=-0.76, P>0.05). Histologic scores of wounds of mice in Huanglian ointment group on PID 7 and 14 were (7.0±1.6) and (11.6±2.1) points respectively, significantly higher than those of vehicle group [(4.2±1.3) and (7.2±1.3) points, with t values respectively 1.96 and 2.50, P<0.05 or P<0.01]. Histologic scores of wounds of mice in two groups on PID 7 and 14 were significantly higher than those on the previous time points of the same group (with P values below 0.01). (3) Percentages of double positive cells of α-SMA and Ki-67 in tissue of wounds of mice in Huanglian ointment group on PID 3 and 7 were (35±12)% and (62±10)% respectively, significantly higher than those of vehicle group [(17±12)% and (34±6)%, with t values respectively -2.48 and -5.25, P<0.05 or P<0.01]. The percentage of double positive cells of α-SMA and Ki-67 in tissue of wounds of mice in Huanglian ointment group on PID 14 was (25±5)%, significantly lower than that of vehicle group [(44±17)%, t=2.50, P<0.05]. The percentage of double positive cells of α-SMA and Ki-67 in tissue of wounds of mice on PID 7 was significantly higher than that on PID 3 or 14 in Huanglian ointment group (with P values below 0.01). Percentages of double positive cells of α-SMA and Ki-67 in tissue of wounds of mice on PID 7 and 14 were significantly higher than those on the previous time points in vehicle group (with P values below 0.05). (4) Protein expressions of TGF-ß in tissue of wounds of mice in Huanglian ointment group on PID 3 and 7 were (396±45) and (722±96) pg/mL respectively, significantly higher than those of vehicle group [(290±42) and (382±62) pg/mL, with t values respectively -8.17 and -6.65, P values below 0.01]. Protein expressions of TGF-ß in tissue of wounds of mice in two groups were close on PID 14 (t=1.60, P>0.05). The protein expression of TGF-ß in tissue of wounds of mice in Huanglian ointment group on PID 7 was significantly higher than that on PID 3 or 14 (with P values below 0.01). Protein expressions of TGF-ß in tissue of wounds of mice in vehicle group on PID 7 and 14 were significantly higher than those on the previous time points (with P values below 0.05). Protein expressions of collagen in tissue of wounds of mice in two groups were close on PID 3 (t=1.99, P>0.05). Protein expressions of collagen in tissue of wounds of mice in Huanglian ointment on PID 7 and 14 were (47±10) and (70±14) ng/mL respectively, significantly higher than those of vehicle group [(34±10) and (42±12) ng/mL, with t values respectively 3.15 and 3.52, P<0.05 or P<0.01]. Protein expressions of collagen in tissue of wounds of mice in two groups on PID 7 and 14 were significantly higher than those on the previous time points of the same group (P<0.05 or P<0.01). Conclusions: Huanglian ointment can promote wound healing of full-thickness skin defect of mice through increasing production of myofibroblasts and protein expressions of TGF-ß and collagen.

Superfamily of genes encoding G protein-coupled receptors in the diamondback moth Plutella xylostella (Lepidoptera: Plutellidae).

G protein-coupled receptors (GPCRs) are the largest and most versatile superfamily of cell membrane proteins, which mediate various physiological processes including reproduction, development and behaviour. The diamondback moth, Plutella xylostella (Lepidoptera: Plutellidae), is one of the most notorious insect pests, preferentially feeding on cruciferous plants. P. xylostella is not only one of the world's most widespread lepidopteran insects, but has also developed resistance to nearly all classes of insecticides. Although the mechanisms of insecticide resistance have been studied extensively in many insect species, few investigations have been carried out on GPCRs in P. xylostella. In the present study, we identified 95 putative GPCRs in the P. xylostella genome. The identified GPCRs were compared with their homologues in Bombyx mori and Drosophila melanogaster. Our results suggest that GPCRs in different insect species may have evolved by a birth-and-death process. One of the differences among compared insects is the duplication of short neuropeptide F receptor and adipokinetic hormone receptors in P. xylostella and B. mori. Another divergence is the decrease in quantity and diversity of the stress-tolerance gene, Mth, in P. xylostella. The evolution by the birth-and-death process is probably involved in adaptation to the feeding behaviour, reproduction and stress responses of P. xylostella. Some of the genes identified in the present study could be potential targets for the development of novel pesticides.

Study on serum proteomic features in patients with and without recurrence or metastasis after surgical resection of esophageal carcinoma.

The purpose of this study was to identify specific bio-markers for recurrence or metastasis of esophageal carcinoma in serum of patients subjected to esophagectomy. Surface-enhanced laser desorp-tion/ionization time-of-flight mass spectrometry (SELDI-TOF-MS) combined with IMAC-Cu(2+) ProteinChip array were performed for the serum protein profiling in patients after surgical resection of esophageal carcinoma. Two groups of patients were analyzed: 38 patients without recurrence or metastasis (Group 1) and 22 patients with recurrence or metastasis after resection (Group 2). The Biomarker Wizard and Bio-marker Patterns software were used to identify proteins differentially expressed between the 2 groups. There were 33 differentially expressed serum proteins detected by comparison between the groups. The clas-sification tree model composed of 3 differentially expressed proteins with different m/z (9368.63, 5342.59, and 5254.43 Da) was established. Under the learning mode, the sensitivity and specificity of this model for diagnosis of esophageal carcinoma recurrence or metastasis were both 100% (22/22 and 38/38, respectively). Under the testing mode, the sensitivity and specificity were 90.9% (20/22) and 94.7% (36/38), re-spectively. The recurrence or metastasis of esophageal carcinoma after esophagectomy can be rapidly and accurately detected using the combi-nation of SELDI-TOF-MS with IMAC-Cu(2+) ProteinChip array, which, therefore, has a potential for clinical application.

A prospective randomized study comparing open versus laparoscopy-assisted D2 radical gastrectomy in advanced gastric cancer.

BACKGROUND: In recent years, many clinical studies have confirmed the value of laparoscopy-assisted gastrectomy (LAG) in gastric cancer surgery, especially in early stages. But the safety and oncologic adequacy of laparoscopy-assisted D2 radical gastrectomy for advanced gastric cancer are still in debate. We conducted a prospective randomized trial to compare open versus laparoscopy-assisted D2 radical gastrectomy in advanced gastric cancer. METHODS: For this study, 123 patients who had been diagnosed endoscopically with gastric cancer were randomly assigned to either LAG (n = 61) or open gastrectomy (OG) (n = 62) which ran from March 2008 to December 2009. Clinical characteristics, operative findings, postoperative recovery, morbidity, pathological report and survival rate were compared. D2 lymph node dissection was performed in 49 patients in the LAG group and 47 patients in the OG group with advanced gastric cancer. We adopt sub-group analysis in this paper. RESULTS: The clinical characteristics of patients in the LAG and OG groups who were in the advanced stage, included age, sex, BMI and concurrent illness, and their ECOG scores were well matched. Operative findings, postoperative recovery, morbidity, pathological findings including tumor location, depth of invasion, TNM stage, histological grade and surgical extension in the two groups were also similar. Compared to the OG group, the mean operating time was significantly longer for the LAG group (267.88 ± 54.284 min in the LAG group vs. 182.02 ± 41.016 min in the OG group, p = 6.383 × 10(-13)); the mean number of days when body temperature exceeded 37°C was significantly shorter in the LAG group (p = 6.34 × 10(-8)). There were no postoperative deaths in both the groups. The postoperative morbidity rate was 12.24% in the LAG group and 19.15% in the OG group with no significant difference (p = 0.357). However, pulmonary infection was observed more frequently in the OG group (p = 0.038). After a mean follow-up of 22.1354 months (from 4 to 36 months), 14 and 15 patients died of gastric cancer in the LAG and OG groups, respectively. Two and one patient died of nongastric cancer in the LAG and OG groups, respectively. The overall survival rates were 67.1% and 53.8% in the LAG and OG groups, respectively. The estimated mean survival time was 29.387 months in the LAG group and 28.978 months in the OG group. There was no statistically significant difference in the overall survival rate for patients in both groups - LAG and OG (log-rank test, p = 0.911, Tarone Ware test, p = 0.994, and Breslow test, p = 0. 961). CONCLUSION: LAG with D2 lymph node dissection is a safe and feasible procedure with adequate lymphadenectomy, good curability and survival rate for the treatment of advanced gastric cancer.

-509C>T polymorphism in the TGF-ß1 gene promoter is not associated with susceptibility to and progression of colorectal cancer in Chinese.

AIM: Colorectal cancer is common, accounting for nearly 10% of all cancers. Transforming growth factor-ß1 (TGF-ß1) is a pleiotropic cytokine that has been implicated in the pathogenesis of colorectal neoplasia. The most studied -509C>T polymorphism of TGF-ß1 gene has been associated with various kinds of cancer. This study investigated the association between this genetic variant and the risk and/or progression of colorectal cancer. METHOD: A case-control study was carried out of 150 colorectal cancer cases and 503 healthy controls. DNA was extracted from blood cell nuclear materials, and -509C>T polymorphism in the TGF-ß1 gene promoter was genotyped by polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP). Colorectal cancer tissues (n = 70) were obtained from the studied cases for measurement of TGF-ß1 mRNA expression levels. We also assessed the plasma TGF-ß1 levels of cases (n = 88) and healthy subjects (n = 120). RESULTS: The TGF-ß1 producer genotype, -509TT, was not associated with an increased risk of colorectal cancer compared with other genotypes. Colorectal cancer patients especially those with a more aggressive disease behaviour were more frequently associated with C allele. CONCLUSION: The results suggest that TGF-ß1 -509C>T polymorphism is not associated with either an increased risk or progression of colorectal cancer.

The expression and antigenicity identification of recombinant rat TGF-beta1 in bacteria.

In order to study structure-function details of TGF-beta1, the recombinant mature form of rat TGF-beta1 was expressed in bacteria. Synthesis of the 112 amino-acid carboxyl-terminal part of TGF-beta1 (amino acid 279-390) was controlled by an inducible gene expression system based on bacteriophage T7 RNA polymerase. This system allowed an active and selective synthesis of recombinant TGF-beta1. The molecular weight of expressed TGF-alpha1 monomer determined on SDS-polyacrylamide gel under reducing conditions was about 13 kD. Serial detergent washes combined with a single gel-filtration purification step were sufficient to purify the expression product to homogeneity. Amino-terminal sequencing revealed that the N-terminal of the recombinant protein was identical to the published data. In Western blot analysis the recombinant polypeptide showed excellent antigenicity against polyclonal TGF-beta1 antibody. The mature recombinant rat TGF-beta1 expressed in this study provides a useful tool for future detailed structural and functional studies.

Caspase-dependent cytosolic release of cytochrome c and membrane translocation of Bax in p53-induced apoptosis.

Activation of p53 induces apoptosis in various cell types. However, the mechanism by which p53 induces apoptosis is still unclear. We reported previously that the activation of a temperature-sensitive mutant p53 (p53(138Val)) induced activation of caspase 3 and apoptosis in Jurkat cells. To elucidate the pathway linking p53 and downstream caspases, we examined the activation of caspases 8 and 9 in apoptotic cells. The results showed that both caspases were activated during apoptosis as judged by the appearance of cleavage products from procaspases and the caspase activities to cleave specific fluorogenic substrates. The significant inhibition of apoptosis by a tetrapeptide inhibitor of caspase 8 and caspase 9 suggested that both caspases are required for apoptosis induction. In addition, the membrane translocation of Bax and cytosolic release of cytochrome c, but not loss of mitochondrial membrane potential, were detected at an early stage of apoptosis. Moreover, Bax translocation, cytochrome c release, and caspase 9 activation were blocked by the broad-spectrum caspase inhibitor, Z-VAD-fmk and the caspase 8-preferential inhibitor, Ac-IETD-CHO, suggesting that the mitochondria might participate in apoptosis by amplifying the upstream death signals. In conclusion, our results indicated that activation of caspase 8 or other caspase(s) by p53 triggered the membrane translocation of Bax and cytosolic release of cytochrome c, which might amplify the apoptotic signal by activating caspase 9 and its downstream caspases.

Construction and selection of the natural immune Fab antibody phage display library from patients with colorectal cancer.

AIM: To construct the natural immune Fab antibody phage display libraries of colorectal cancer and to select antibodies related with colorectal cancer. METHODS: Extract total RNA from tissue of local cancer metastasis lymph nodes of patients with colorectal cancer. RT-PCR was used to amplify the heavy chain Fd and light chain kappa and the amplification products were inserted successively into the vector pComb3 to construct the human libraries of Fab antibodies. They were then panned by phage display technology. By means of Dot immunoblotting and ELISA, the libraries were identified and the Fab phage antibodies binding with antigens of colorectal cancer were selected. RESULTS: The amplified fragments of Fd and kappa gained by RT-PCR were about 650 bp. Fd and kappa PCR products were subsequently inserted into the vector pComb3, resulting in a recombination rate of 40% and the volume of Fab phage display library reached 1.48 x 10(6). The libraries were enriched about 120-fold by 3 cycles of adsorption-elution-multiplication (panning). Dot immunoblotting showed Fab expressions on the phage libraries and ELISA showed 5 clones of Fab phage antibodies which had binding activities with antigens of colorectal cancer. CONCLUSION: The natural immune Fab antibody phage display libraries of colorectal cancer were constructed. They could be used to select the relative antibodies of colorectal cancer.