Discovering Innate Driver Variants for Risk Assessment of Early Colorectal Cancer Metastasis.

Metastasis is the main fatal cause of colorectal cancer (CRC). Although enormous efforts have been made to date to identify biomarkers associated with metastasis, there is still a huge gap to translate these efforts into effective clinical applications due to the poor consistency of biomarkers in dealing with the genetic heterogeneity of CRCs. In this study, a small cohort of eight CRC patients was recruited, from whom we collected cancer, paracancer, and normal tissues simultaneously and performed whole-exome sequencing. Given the exomes, a novel statistical parameter LIP was introduced to quantitatively measure the local invasion power for every somatic and germline mutation, whereby we affirmed that the innate germline mutations instead of somatic mutations might serve as the major driving force in promoting local invasion. Furthermore, via bioinformatic analyses of big data derived from the public zone, we identified ten potential driver variants that likely urged the local invasion of tumor cells into nearby tissue. Of them, six corresponding genes were new to CRC metastasis. In addition, a metastasis resister variant was also identified. Based on these eleven variants, we constructed a logistic regression model for rapid risk assessment of early metastasis, which was also deployed as an online server, AmetaRisk (http://www.bio-add.org/AmetaRisk). In summary, we made a valuable attempt in this study to exome-wide explore the genetic driving force to local invasion, which provides new insights into the mechanistic understanding of metastasis. Furthermore, the risk assessment model can assist in prioritizing therapeutic regimens in clinics and discovering new drug targets, and thus substantially increase the survival rate of CRC patients.

Homocysteine, hypertension, and risks of cardiovascular events and all-cause death in the Chinese elderly population: a prospective study.

BACKGROUND: Increased homocysteine levels are associated with the risk of cardiovascular disease (CVD) and death. However, their prevention has not been effective in decreasing CVD risk. This study investigated the individual and combined associations of hyperhomocysteinemia and hypertension with incident CVD events and all-cause death in the Chinese elderly population without a history of CVD. METHODS: This prospective study was conducted among 1,257 elderly participants (mean age: 69 years). A questionnaire survey, physical examinations, and laboratory tests were conducted to collect baseline data. Hyperhomocysteinemia was defined as homocysteine level ≥ 15 µmol/L. H-type hypertension was defined as concomitant hypertension and hyperhomocysteinemia. Multivariate Cox regression analysis was used to evaluate individual and combined associations of hyperhomocysteinemia and hypertension with the risks of incident CVD events and all-cause death. RESULTS: Over a median of 4.84-year follow-up, hyperhomocysteinemia was independently associated with incident CVD events and all-cause death. The hazard ratios (HRs) were 1.45 (95% CI: 1.01-2.08) for incident CVD events and 1.55 (95% CI: 1.04-2.30) for all-cause death. After adjustment for confounding factors, H-type hypertension had the highest HRs for incident CVD events and all-cause death. The fully adjusted HRs were 2.44 for incident CVD events (95% CI: 1.28-4.65), 2.07 for stroke events (95% CI: 1.01-4.29), 8.33 for coronary events (95% CI: 1.10-63.11), and 2.31 for all-cause death (95% CI: 1.15-4.62). CONCLUSIONS: Hyperhomocysteinemia was an independent risk factor, and when accompanied by hypertension, it contributed to incident CVD events and all-cause death in the Chinese elderly population without a history of CVD.

Listeria monocytogenes bacteremia in a centenarian and pathogen traceability: A case report.

BACKGROUND: Early diagnosis and appropriate antibiotic treatment are important to survival of Listeria monocytogenes (L. monocytogenes) bacteremia. Penicillin tends to be the most commonly used antibiotic. However, there are limited data on antibiotic use in elderly patients with serious complications. We describe the clinical presentation, antibiotic therapy, and traceability of L. monocytogenes in a centenarian with a history of eating frozen food. CASE SUMMARY: A 102-year-old man suffered from high fever with chill after hematochezia. Tentative diagnoses were lower gastrointestinal hemorrhage and localized peritonitis. Meropenem and ornidazole were the empirical therapy. The patient did not respond and developed multiple system dysfunction even after teicoplanin was added to the therapy. L. monocytogenes was identified from blood cultures on day 5 of admission. The patient had a history of consuming frozen dumplings. Meropenem/ornidazole/teicoplanin were replaced with meropenem/linezolid. The patient gradually became afebrile. He received meropenem/linezolid for 10 d, and piperacillin/tazobactam was applied as step-down treatment for 2 wk with good clinical results. There was no sign of relapse during follow-up after discharge. L. monocytogenes isolates from the patient and frozen dumplings belonged to different serotypes and sequence types (STs): 1/2b and ST5 from the patient and 1/2c and ST9 from the dumplings. CONCLUSION: More awareness of listeriosis should be raised. Linezolid might be an option for listeriosis in elderly people with serious complications.

Silencing of DHX32 increases the proliferation of liver cancer cells.

BACKGROUND: Liver cancer ranks fifth in malignancy incidence globally and is the second leading cause of cancer-related death in China. Chronic hepatitis B or C infection and alcohol abuse have been identified to be the major risk factors for liver cancer development. Some evidence implicates DHX32 as being critically involved in tumor progression. The role of DHX32 in liver cancer specifically, however, remains unclear. METHODS: Fifty-three liver cancer tissue and paracancerous tissue samples were surgically resected from 53 patients who were admitted to Zhongshan Hospital between 2006 and 2008. We used immunohistochemistry (IHC) to analyze the expressions of DHX32, established liver cancer cells with stable DHX32 knockdown, and investigated the proliferation of these cells with methyl thiazolyl tetrazolium (MTT) and 5-ethynyl-2'-deoxyuridine (EdU) data. RESULT: Baseline characteristics of enrolled liver cancer patients (53 patients) were summarized, and the IHC results firstly showed that 88.7% (47/53) of paracancerous tissues exhibited a high expression of DHX32, while only 43.4% (23/53) of liver cancer tissues showed similar expression. We then established liver cancer cells with the stable knockdown of DHX32. MTT and EdU data demonstrated that DHX32 knockdown in liver cancer cells enhanced the proliferative potential of liver cancer cells. Furthermore, phosphorylated levels of extracellular signal-regulated kinase (ERK) and protein kinase B (Akt) were upregulated in liver cancer cells with DHX32 knockdown. We also found the level of cyclin-dependent kinases 6 (CDK6) to be increased in liver cancer cells with DHX32 knockdown. CONCLUSIONS: DHX32 showed a lower expression in liver cancer tissues than in paracancerous tissues and could harbor a proliferation-suppressing property in liver cancer. DHX32 may thus be a possible target for gene therapy.

Performance evaluation of a chemiluminescence microparticle immunoassay for CK-MB.

BACKGROUND: To verify and evaluate the performance characteristics of a creatine kinase phosphokinase isoenzymes MB (CK-MB) assay kit, which produced by Xiamen Innodx Biotech Co. Ltd. METHODS: Evaluation was carried out according to "Guidelines for principle of analysis performance evaluation of in vitro diagnostic reagent." The performance parameters included detection limit, linearity range, reportable range, recovery test, precision verification, interference test, cross-reactivity, matrix effect, and method comparison. RESULTS: The detection limit was 0.1 ng/mL. The assay had clinical linearity over range of 0.1 ng/mL-500 ng/mL. Reportable range was from 0.1 ng/mL to 1000 ng/mL. The average percent of recovery was 99.66%. The coefficient of variation (CV) for within-run and between-run of low CK-MB sample was 5.55% and 6.16%, respectively. As for high-level sample, it was 7.88% and 7.80%. In medical decision level, the relative deviation (Bias) of all interference tests was lower than 15%. When the sample had mild-hemolysis; hemoglobin ≤15 g/L; triglyceride ≤17 mmol/L; bilirubin ≤427.5 µmol/L; rheumatoid factor ≤206U/mL, there was no significant interference to be found. Moreover, assay kit had no cross-reaction with CK-MM and CK-BB. At last, total diagnostic accuracy of kit was 93.24%, when compared with refer kit. CONCLUSION: Overall the results of the verification study indicated the performance of kit is met the requirements of the clinical test.

Evaluation of a newly developed chemiluminescence immunoassay for detecting cardiac troponin T.

BACKGROUND: To evaluate the performance of a chemiluminescence detection kit for cardiac troponin T (cTnT). METHODS: According to the "Guiding principles on performance analysis of diagnostic reagents in vitro" and the Clinical and Laboratory Standards Institute (CLSI) Guidelines, we assessed the detection limit, linear range, reportable range, accuracy, precision, cross reactivity, interference factors, and matrix effect of the kit, and compared these parameters with that of the commercial electrochemiluminescence detection kit for cTnT (Roche). RESULTS: The minimum detection limit of the kit was 0.01 ng/mL. The linear range was 0.01 ng/mL-25 ng/mL. The reportable range was from 0.01 ng/mL to 100 ng/mL. Precision within the batch was 2.9%-6.4%, and precision between batches was 6.0%; the accuracy was good and the recovery rate reached 96.2%. The cross-reaction test demonstrated that there was no reactivity between cTnT and a variety of troponins. Test results deviated by less than ±10% in the presence of hemoglobin ≤1000 µg/mL, bilirubin ≤250 µg/mL, triglycerides ≤11.25 mg/mL, and rheumatoid factor ≤206 U/mL, suggesting that kit results were not significantly interfered with these factors. Results from the matrix-effect assessment revealed absence of a matrix effect in the tested serum samples. Correlation study revealed that the performance of the kit was very consistent with that of the Roche electrochemiluminescence detection kit (Kappa = 0.900, P < .001) with a high correlation (r = .903, P < .001) and a total matching rate of 95.00%. CONCLUSION: The performance of the evaluated chemiluminescence immunoquantitation kit for cTnT detection was acceptable in all tested parameters, fulfilling requirements for clinical applications.

Identification of two novel genes SLC15A2 and SLCO1B3 associated with maintenance dose variability of warfarin in a Chinese population.

Warfarin is a commonly prescribed and effective oral anticoagulant. Genetic polymorphisms associated with warfarin metabolism and sensitivity have been implicated in the wide inter-individual dose variation that is observed. Several algorithms integrating patients' clinical characteristics and genetic polymorphism information have been explored to predict warfarin dose. However, most of these algorithms could explain only over half of the variation in a warfarin maintenance dose, suggesting that additional genetic factors may exist and need to be identified. Here, a drug absorption, distribution, metabolism and excretion (ADME) Core Panel Kit-based pharmacogenetic study was performed to screen for warfarin dose-associated SNP sites in Han-Chinese population patients taking warfarin therapy, and the screen was followed by pyrosequencing-based validation. Finally, we confirmed that the common variant rs9923231 in VKORC1 and two novel genes, SLC15A2 (rs1143671 and rs1143672) and SLCO1B3 (rs4149117 and rs7311358), are associated with the warfarin maintenance dose. As has been shown for those carriers with the variant rs9923231 in VKORC1, it was suggested that those subjects with homozygous minor alleles in those four SNPs should take a lower warfarin dose than those carrying the wild type alleles. Together with the established predictor rs9923231 in VKORC1, those four novel variants on SLC15A2 and SLCO1B3 should be considered as useful biomarkers for warfarin dose adjustment in clinical practice in Han-Chinese populations.

Plasma microRNA-19a as a potential biomarker for esophageal squamous cell carcinoma diagnosis and prognosis.

AIM: To investigate whether plasma miR-19a can serve as a biomarker for esophageal squamous cell carcinoma (ESCC) diagnosis and prognosis. MATERIALS & METHODS: Plasma samples from 89 ESCC, 45 benign lesion patients and 80 healthy controls were subjected to RT-qPCR analyses for miR-19a. In addition, plasma samples from 30 patients were collected before and after surgery for the same analyses. RESULTS: Plasma miR-19a was significantly increased in ESCC patients compared with healthy controls. The sensitivity of miR-19a for early stages of ESCC was 68.09%. Combination of miR-19a and cytokeratin 19 fragment 21-1 (Cyfra21-1) further improved the sensitivity to 78.70%. Moreover, plasma miR-19a level was decreased in patients after surgery. CONCLUSION: Plasma miR-19a may serve as a potential biomarker that complements Cyfra21-1 in detecting early stages of ESCC.

DHX32 Promotes Angiogenesis in Colorectal Cancer Through Augmenting ß-catenin Signaling to Induce Expression of VEGFA.

We previously reported that overexpression of DHX32 contributes to the growth and metastasis of colorectal cancer (CRC). However, the underlying mechanism is not largely characterized. Herein, we reported that DHX32 in CRC cells upregulated expression of vascular endothelial growth factor A (VEGFA) at the transcription level through interacting with and stabilizing ß-catenin. This promoted the recruitment of host endothelial cells to the tumor, and therefore, formation of microvessel in the tumor. Xenograft model revealed that depletion of DHX32 in CRC cells significantly reduced the microvessel density in the grafts and suppressed the growth of grafts. Furthermore, the expression level of DHX32 was positively associated with microvessel density in human CRC and poor outcome of CRC patients. Therefore, the report demonstrates that DHX32 is a pro-angiogenic factor, that inhibition of DHX32-ß-catenin pathway can provide a strategy for CRC treatment, and that the expression level of DHX32 has the potential to serve as a biomarker for CRC diagnosis and prognosis.

Depletion of Cks1 and Cks2 expression compromises cell proliferation and enhance chemotherapy-induced apoptosis in HepG2 cells.

The present study explored the oncogenic roles of overexpressed Cks1 and Cks2 in human hepatocellular carcinoma cells. Gene expression of Cks1 and Cks2 in HepG2 cells was disrupted by siRNA or increased by cDNA transfection. Cell proliferation was assayed by CCK-8 analysis and cell counting. Cisplatin-induced apoptosis after transfection was measured by flow cytometry using Annexin V/propidium iodide (PI) double staining. Cell cycle changes after transfection were determined by flow cytometry with PI staining. Protein levels of Akt and GSK-3ß were measured after transfection. The results revealed that HepG2 proliferation was decreased by depletion of endogenous Cks1 or Cks2, and increased by overexpression of Cks1 or Cks2. HepG2 apoptosis increased concordantly with the decline of Cks1 or Cks2 expression. Overexpression of Cks1 or Cks2 prevented cell apoptosis. Protein levels of p­Akt and p­GSK-3ß were downregulated after RNA interference of Cks1 or Cks2. In conclusion, Cks1 and Cks2 promoted proliferation and prevented apoptosis of HepG2 cells. The Akt/GSK-3ß-related PI3K/Akt signaling pathway may be a key signaling pathway that is involved in the regulation of cell growth and cell death.

Plasma levels of microRNA-24, microRNA-320a, and microRNA-423-5p are potential biomarkers for colorectal carcinoma.

BACKGROUND: MicroRNAs are stable and easy to detect in plasma. The plasma levels of microRNAs are often changed in disease conditions, including cancer. This makes circulating microRNAs a novel class of biomarkers for cancer diagnosis. Analyses of online microRNA data base revealed that expression level of three microRNAs, microRNA-24 (miR-24), microRNA-320a (miR-320a), and microRNA-423-5p (miR-423-5p) were down-regulated in colorectal cancer (CRC). However, whether the plasma level of these three microRNAs can serve as biomarkers for CRC diagnosis and prognosis is not determined. METHODS: Plasma samples from 223 patients with colorectal related diseases (111 cancer carcinoma, 59 adenoma, 24 colorectal polyps and 29 inflammatory bowel disease) and 130 healthy controls were collected and subjected to reverse transcription-quantitative real time PCR (RT-qPCR) analyses for the three microRNAs. In addition, plasma samples from 43 patients were collected before and after surgical treatment for the same RT-qPCR analyses. RESULTS: The concentrations of plasma miR-24, miR-320a and miR-423-5p were all decreased in patients with CRC and benign lesions (polyps and adenoma) compared with healthy controls, but increased in inflammatory bowel disease (IBD). The sensitivity of miR-24, miR-320a and miR-423-5p for early stage of CRC were 77.78 %, 90.74 %, and 88.89 %, respectively. Moreover, the plasma concentration of the three microRNAs was increased in patients after the surgery who had clinical improvement. CONCLUSIONS: The plasma levels of miR-24, miR-320a, and miR-423-5p have promising potential to serve as novel biomarkers for CRC detection, especially for early stage of CRC, which are superior to the currently used clinical biomarkers for CRC detection, such as CEA and CA19-9. Further efforts to develop the three microRNAs as biomarkers for early CRC diagnosis and prediction of surgical treatment outcomes are warrant.

Overexpression of DHX32 contributes to the growth and metastasis of colorectal cancer.

Our previous work demonstrates that DHX32 is upregulated in colorectal cancer (CRC) compared to its adjacent normal tissues. However, how overexpressed DHX32 contributes to CRC remains largely unknown. In this study, we reported that DHX32 was overexpressed in human colon cancer cells. Overexpressed DHX32 promoted SW480 cancer cells proliferation, migration, and invasion, as well as decreased the susceptibility to chemotherapy agent 5-Fluorouracil. Furthermore, PCR array analyses revealed that depleting DHX32 in SW480 colon cancer cells suppressed expression of WISP1, MMP7 and VEGFA in the Wnt pathway, and anti-apoptotic gene BCL2 and CA9, however, elevated expression of pro-apoptotic gene ACSL5. The findings suggested that overexpressed DHX32 played an important role in CRC progression and metastasis and that DHX32 has the potential to serve as a biomarker and a novel therapeutic target for CRC.

Overexpression of DEK gene is correlated with poor prognosis in hepatocellular carcinoma.

The oncogene DEK was originally identified as one of the parts of the DEK­CAN fusion gene, arising from the translocation (6;9) in a subtype of acute myeloid leukemia. Since then, DEK has been shown to promote tumorigenesis in a variety of cancer cell types through its roles in inhibiting cell differentiation, senescence and apoptosis. Certain studies have established that DEK is dysregulated in several types of cancer, including hepatocellular carcinoma (HCC). However, its clinical significance in human HCC remains unknown. In this study, the expression of DEK mRNA and protein was examined in 55 surgical HCC specimens and matched non­tumorous tissues. In addition, the correlation between DEK expression and clinicopathological characteristics and prognosis was analyzed. mRNA and protein levels of DEK were found to be significantly overexpressed in the majority of HCC tumors when compared with matched normal hepatic tissues (P<0.05). In addition, the expression pattern of DEK was closely correlated with differentiation status, portal venous invasion and tumor size (P<0.05). Kaplan­Meier curves demonstrated that patients with higher DEK expression levels had significantly poorer survival than those with lower DEK expression levels (P=0.003). In addition, Cox regression analysis demonstrated that the level of DEK expression may be a valuable prognostic factor (P<0.05). These results suggested that DEK may play a significant role in hepatocyte differentiation and may serve as a useful prognostic marker and biomarker for the staging of HCC.

Clinical significance of combined testing of YKL-40 with CEA in Chinese colorectal cancer patients.

BACKGROUND: To investigate the practical value of individual and combined testing of plasma levels of YKL-40, CEA, and CA199 for auxiliary diagnosis and detection of recurrence of colorectal cancer. METHODS: ELISA and ECLIA were used to evaluate levels of YKL-40, CEA, and CA199 in 120 colorectal cancer patients (56 initial-diagnosis, 42 post-operative, and 22 recurrent cases). Forty-three patients with benign colorectal disease and 36 healthy patients were enrolled as controls. The relationship between YKL-40 and clinical indicators of tumor pathology was analyzed. The positive rate and diagnostic efficacy of single and combined YKL-40, CEA, and CA199 testing were assessed in patients with colorectal cancer. RESULTS: Plasma YKL-40 in the cancer group was significantly higher than in the benign control and healthy control group, and the mean values were 145.4 ng/mL, 107.7 ng/mL, and 51.3 ng/mL (p < 0.05), respectively. With 72 ng/mL as the diagnostic threshold, the sensitivity and specificity of YKL-40 in colorectal cancer diagnosis were found to be 73.2% and 66.7%, respectively. Early-stage colorectal cancer patients showed a YKL-40 positive rate (73%) significantly higher than those of CEA and CA199 (50% and 32%, respectively; p < 0.05). When YKL-40 testing was combined with CEA or CA199, the positive rate increased to 82.1% and 80.3%, respectively. Through ROC curve analysis of the post-operative recurrent group against the non-recurrent group, the areas under the curve for YKL-40, CEA, and CA199 were found to be 0.907, 0.714, and 0.759, respectively. Based on the Dukes classification, the mean YKL-40 value for stages A/B, C, and D were 120.1 ng/mL, 131.7 ng/mL, and 226.8 ng/mL (p = 0.008), respectively. The plasma YKL-40 level gradually increased as the disease progressed. Lower degrees of tumor differentiation were correlated with higher YKL-40 levels. The mean YKL-40 values of high, medium, and low tumor differentiation groups were 96.8 ng/mL, 127.5 ng/mL, and 225.7 ng/mL (p = 0.004), respectively. CONCLUSIONS: The benefits of using YKL-40 testing are higher than CEA and CA199 for the monitoring of colorectal cancer recurrence. Combined testing of both YKL-40 and CEA was found to be optimal for auxiliary diagnosis of colorectal cancer. Plasma YKL-40 was found to be suitable for auxiliary diagnosis of colorectal cancer.

Characteristics of patients suffering from cow milk allergy.

The most frequent symptoms among the manifestations of cow milk allergy (CMA) are gastrointestinal. CMA pathogenesis involves immunological mechanisms with participation of immunocompetent cells, production of immunoglobulin E (IgE) and immunoglobulin G (IgG). We aim to determine whether cow milk-specific IgE antibodies coexist with cow milk-specific IgG antibodies in CMA patients with diarrhea symptom, and if there is any relationship between both antibody types. 65 CMA patients (average age of 17 years, ranging from 2 to 74 years), all of who had diarrhea symptom of CMA, were enrolled in this study. The total cow IgE and IgG subclass in serum were measured by electrochemiluminescence immunoassay and rate immune scatter turbidimetry, respectively. And also the cow milk-specific IgE was determined by enzyme-linked immunosorbent assay. The number of eosinophils in serum was calculated by Sysmex XE-2100 Hematology Analyzer. Our data showed that both cow milk-specific IgG and IgE levels were significantly elevated in CMA patients compared to those of age-matched control subjects. Out of the 65 CMA patients, 40 showed elevated cow milk-specific IgE antibody level, among which, 28 cases presented highly sensitive reaction to cow milk-specific IgG, along with each six of moderate and mild sensitive reaction to cow milk-specific IgG; while 20 showed elevated total IgG levels. The IgG3 positive rate was 16.9%, which was the highest. A moderate correlation between cow milk-specific IgE and cow milk-specific IgG was found in the CMA patients (r=0.415, P=0.001). The results indicated that cow milk-specific IgE antibodies could coexist with cow milk-specific IgG antibodies in patients suffering from CMA. The aberrant changes in the concentration of cow milk-specific IgE antibodies were associated with cow milk-specific IgG antibodies.

Effects of silencing cyclooxygenase-2 expression via RNA interference on the tumorigenicity of the SMMC-7721 human hepatocarcinoma cell line.

We constructed a vector carrying a shRNA sequence against cyclooxygenase-2 (COX-2) that was subsequently transfected into the human hepatocarcinoma cell line SMMC­7721. Furthermore, we established a COX-2-deficient stable cell line and a model of tumor-shRNA transplantation in nude mice. Negative shRNA was used as the control. The tumor volume in the experimental group was smaller compared to that in the control group. Hematoxylin and eosin staining indicated that the cells in the experimental group differentiated better than those in the control group. The COX-2 mRNA level in the tumor tissues injected with SMMC-7721/COX-2i was markedly downregulated compared to that in the tumor tissues injected with SMMC-7721/negative shRNA. The inhibition rate reached 68.6%. Immunohistological study showed a significantly strong COX-2 expression in the control group tumor cells, whereas the experimental group exhibited moderate expression, indicating the inhibition of COX-2 expression after transfection of cells with shRNA against COX-2. Western blot analysis further proved the inhibition of COX-2 expression. In conclusion, RNAi-mediated regulation of COX-2 expression could efficiently inhibit liver-transplanted tumor growth in BALB/c nude mice.

Expression of inflammatory and apoptosis factors following coronary stent implantation in coronary heart disease patients.

We investigated the changes in characteristics of neutrophil CD11b, monocyte CD11b, platelet CD62P, endothelin (ET), and neutrophil CD178 in patients with coronary heart disease (CHD) before and after primary coronary stenting. A total of 41 patients with CHD who underwent coronary stenting and 40 control subjects were enrolled in the study. In CHD patients, peripheral blood samples were taken 24 h before and 30 min, 24 h, and 72 h after successful coronary stenting. All markers were significantly elevated in patients with CHD compared with controls (P<0.05). Time-course studies revealed that the expressions of neutrophil CD11b, monocyte CD11b, platelet CD62P, and ET were lower at 30 min post-operation (PO) compared with that at 24 h before operation (BO) (P<0.05). All levels significantly increased from 30 min PO to 24 h PO (P<0.05) and decreased thereafter until 72 h PO (P>0.05). Time course changes in neutrophil CD11b levels after coronary stenting were significantly higher in patients with unstable angina pectoris than in patients with stable angina pectoris (P<0.05). CD11b levels were related to CD62P in patients with CHD (P<0.05). Neutrophil CD11b and monocyte CD11b levels were significantly increased in patients with CHD who underwent coronary stenting compared with controls (P<0.05). Results show that CD11b levels increased, meanwhile, the levels of CD62P and ET increased in CHD patients after coronary stenting. In addition, neutrophil CD178 levels of apoptosis factor in patients, which is important for regression of inflammation, remained high for a period of time after coronary stenting.

Optimization of conditions for transfection with the Sofast gene vector.

We previously reported the synthesis and characterization of a novel cationic polymer gene vector. The present article further explored and optimized the working conditions of the Sofast gene vector both in vitro and in vivo, and improved its performance. The transfection conditions of Sofast, such as cell type, cell density, transfection time, N/P values and analysis time after transfection, were further explored. Moreover, the effects of the fusion peptide diINF-7 on transfection efficiency were examined. Sofast was successfully applied for the transfection of exogenous genes into more than 40 types of cell lines derived from humans, mice, monkeys and other species. When the cells were 50-80% confluent, Sofast possessed a better transfection efficiency. In most cases, Sofast also had a higher transfection efficiency when it was used to transfect cells that were seeded for several hours and had adhered to the substrate. The results from in vitro experiments indicate that the recommended Sofast to DNA mass ratio is 16:1, and the optimum analysis time after transfection is 48 h. The salt concentration in the Sofast working solution markedly affected the transfection efficiency. When conducting in vivo transfection, the working solution should be salt-free, whereas for in vitro transfection, it is more appropriate for the working solution to include certain salt concentrations. Finally, the results confirm that diINF-7 significantly promotes the transfection efficiency of Sofast. In conclusion, the present research not only established the optimal conditions for Sofast in the transfection of commonly used cells, but also built the foundations for in vivo and in vitro applications of Sofast, as well as its use in clinical practice.

Evaluation of a colloidal gold immunochromatography assay in the detection of Treponema pallidum specific IgM antibody in syphilis serofast reaction patients: a serologic marker for the relapse and infection of syphilis.

Syphilis remains as a worldwide public health problem; hence, it is necessary to develop a new diagnostic approach that is easier and faster than conventional tests. A new testing method to detect Treponema pallidum IgM (TP-IgM), named colloidal gold immunochromatography assay (GICA), is presented in place of fluorescent treponemal antibody absorption (FTA-Abs). TP-IgM was detected using GICA developed on syphilis-specific recombinant proteins TPN17 and TPN47. The FTA-Abs IgM test was set as the gold standard. A GICA TP-IgM test was performed to detect syphilis in 1208 patients who received recommended therapy for syphilis for more than 1 year at the Xiamen Center of Clinical Laboratory in China from June 2005 to May 2009. One hundred blood donors were set up as control. The sensitivity, specificity, positive predictive value, negative predictive value, positive likelihood ratio, and negative likelihood ratio were 98.21%, 99.04%, 93.75%, 99.73%, 102.3, and 0.018, respectively. Detection on 500 interference specimens indicated that the biological false-positive rate of the GICA test was extremely low and was free from other biological and chemical factors. The patients were divided into the following experimental groups based on the results of toluidine red unheated serum test (TRUST) and treponemal pallidum particle agglutination (TPPA): (1) the syphilis serofast reaction (SSR) group consisted of 411 cases with (+) TRUST and (+) TPPA, which exhibited no clinical manifestations of syphilis after 1 year of recommended syphilis treatment; (2) the serum cure group, which was further subdivided into group A, a group that consisted of 251 cases with (-) TRUST and (+) TPPA, and (3) group B, a group that consisted of 546 cases with (-) TRUST and (-) TPPA; and (4) the blood donor control group, which consisted of 100 healthy persons with (-) ELISA-TP and (-) TPPA. We used the FTA-Abs method and the GICA method to detect TP-IgM; the positive rate of TP-IgM in 411 SSR patients was 34.55% and 36.01%, respectively. However, in serum cure group A, the positive rate of TP-IgM was 10.36% and 11.16%, respectively. The χ(2) test revealed that there is a significant difference in the positive rate between these 2 groups (P < 0.01). The TP-IgM positive rate in the same group, as detected by the GICA method and the FTA-Abs method, had no significant difference in statistics. However, as detected by the GICA method and the FTA-Abs method, all the samples in serum cure group B and the control group were negative for TP-IgM. The TP-IgM-positive result demonstrated that active T. pallidum remained in the bodies of SSR patients. In summary, the characteristics of GICA TP-IgM correspond to that of FTA-Abs TP-IgM; this can be used as a serologic marker for the relapse and infection of syphilis in place of the conventional FTA-Abs IgM test.

Further evaluation of a novel nano-scale gene vector for in vivo transfection of siRNA.

In this research, a lipid-cationic polymer (LCP) containing the side-chain branching of brassidic acid was synthesized using chemical methods. As a gene vector for small interfering ribonucleic acid (siRNA) transfection, the efficiency and biosafety of LCP were preliminarily evaluated to investigate its possible application on tumor gene therapy. The toxicity, side-effects, and biosafety of LCP were investigated in animals based on the results of in vitro experiments. The siRNA against cyclooxygenase-2 (COX-2) was transfected by LCP to interfere with the COX-2 expression in nude-transplanted tumors. Hematoxylin and eosin stains, immunohistochemistry, reverse transcription-polymerase chain reaction, and Western blot were performed to evaluate the efficiency of LCP for siRNA transfection. The animal toxicity experiment showed that a high concentration of LCP had a low toxic effect on animals and did not induce allergic or pyrogenic reactions. The results from the in vivo transfection indicated that LCP could efficiently transfect siRNA and silence the target gene expression. The LCP gene vector for siRNA transfection is highly efficient during in vivo transfection and had low toxicity. From all aspects of tumor gene therapy and basic research, LCP is valuable for scientific research and medical applications.