Retraction Note: Nuclear factor of activated T cells 5 maintained by Hotair suppression of miR-568 upregulates S100 calcium binding protein A4 to promote breast cancer metastasis.

The authors are retracting this article [1] after an investigation by the Ethics Committee of the Fourth Military Medical University (Xi'an, Shaanxi, China) of the following concerns that had been raised with respect to two of the figures.

Downregulation of DHRS9 expression in colorectal cancer tissues and its prognostic significance.

Dehydrogenase/reductase (SDR family) member 9 (DHRS9) is aberrantly expressed in colorectal cancer (CRC), but its prognostic value is unknown. The aim of the work was to investigate the prognostic significance of DHRS9 expression in CRC. We found that DHRS9 was frequently downregulated in CRC clinical samples at both the messenger RNA (mRNA) and protein levels. Decreased expression of DHRS9 was significantly correlated with increased lymph node metastasis (p = 0.032), advanced tumor-node-metastasis (TNM) stage (p = 0.021), increased disease recurrence (p = 0.001), and death (p = 0.014). Kaplan-Meier analysis indicated that low DHRS9 expression predicted poor disease-free survival (p = 0.003) and disease-specific survival (p = 0.021). Cox multivariate analysis revealed that reduced expression of DHRS9 was an independent unfavorable prognostic indicator for CRC. Furthermore, combination of DHRS9 with TNM stage was a more powerful predictor of poor prognosis than either of the two parameters alone. Our results suggest that decreased expression of DHRS9 correlates with tumor progression and may serve as a potential prognostic biomarker in CRC.

Overexpression of SULT2B1b is an independent prognostic indicator and promotes cell growth and invasion in colorectal carcinoma.

Aberrant expression of cytosolic sulfotransferase 2B1b (SULT2B1b) has been reported in several human malignancies. However, the expression pattern and clinical significance of SULT2B1b in colorectal carcinoma (CRC) remains unknown. Real-time quantitative PCR, western blot, and immunohistochemistry analyses were used to determine SULT2B1b expression in CRC clinical samples and CRC-derived cell lines. Kaplan-Meier and Cox proportional regression analyses were used to evaluate the association between SULT2B1b expression and patient survival in two independent cohorts of 485 patients with CRC. Gain- and loss-of-function approaches were employed to investigate the role of SULT2B1b in regulation of CRC cell growth and invasion. We found that SULT2B1b expression was frequently upregulated in CRC clinical samples and CRC-derived cell lines and was significantly correlated with lymph node metastasis and TNM stage in both the training and validation cohorts. Patients with higher intratumoral SULT2B1b expression had a significantly shorter disease-specific survival (DSS) and disease-free survival (DFS) than those with lower expression. Importantly, increased expression of SULT2B1b significantly predicted poor DSS and DFS and was an independent unfavorable prognostic indicator for stage II patients in both cohorts. Functional studies revealed that overexpression of SULT2B1b promoted CRC cell growth and invasion in vitro. Conversely, knockdown of SULT2B1b inhibited these processes. In conclusion, our findings suggest that SULT2B1b expression correlates with disease progression and metastasis and may serve as a novel prognostic biomarker and potential therapeutic target for patients with CRC.

Multicenter analysis of soluble Axl reveals diagnostic value for very early stage hepatocellular carcinoma.

If diagnosed at early stages, patients with hepatocellular carcinoma (HCC) can receive curative therapies, whereas therapeutic options at later stages are very limited. Here, we addressed the potential of soluble Axl (sAxl) as a biomarker of early HCC by analyzing levels of sAxl in 311 HCC and 237 control serum samples from centers in Europe and China. Serum concentrations of sAxl were significantly increased in HCC (18.575 ng/mL) as compared to healthy (13.388 ng/mL) or cirrhotic (12.169 ng/mL) controls. Receiver operating characteristic curve analysis of sAxl in very early stage HCC patients (BCLC 0) showed an area under the curve (AUC) of 0.848, with a sensitivity of 76.9% and a specificity of 69.2%. α-Fetoprotein (AFP)-negative HCC patients displayed an AUC of 0.803, with sensitivity and specificity of 73% and 70.8%. Combination of sAxl and AFP improved diagnostic accuracy to 0.936 in very early HCC patients and to 0.937 in all HCC. Differential diagnosis of very early HCC versus liver cirrhosis showed a combined performance for sAxl and AFP of 0.901 with a sensitivity of 88.5% and a specificity of 76.7%. Furthermore, sAxl levels failed to be elevated in primary ovarian, colorectal and breast carcinomas as well as in secondary hepatic malignancies derived from colon. In summary, sAxl outperforms AFP in detecting very early HCC as compared to healthy or cirrhotic controls and shows high diagnostic accuracy for AFP-negative patients. sAxl is specific for HCC and suggested as a biomarker for routine clinical use.

Serine threonine tyrosine kinase 1 is a potential prognostic marker in colorectal cancer.

BACKGROUND: Aberrant expression of serine threonine tyrosine kinase 1 (STYK1) has been reported in several human malignancies including colorectal cancer (CRC). However, the prognostic significance of STYK1 expression in CRC remains unknown. METHODS: STYK1 protein expression in paraffin-embedded CRC specimens was determined immunohistochemically. The correlation of STYK1 expression with clinicopathologic features was assessed in a cohort containing 353 patients with primary CRC. Kaplan-Meier and Cox proportional regression analyses were used to evaluate the association between STYK1 expression and patients' survival. RESULTS: STYK1 expression was frequently up-regulated in CRC clinical samples at the protein levels and was significantly associated with tumor differentiation grade (p = 0.030), lymph node metastasis (p = 0.004), TNM stage (p = 0.007) and patient death (p < 0.001). Kaplan-Meier analysis indicated that patients with high intratumoral STYK1 expression had a significantly shorter disease-specific survival (DSS) than those with low expression (p < 0.001). Importantly, high levels of STYK1 protein predicted poor DSS for both stage II (p < 0.001) and stage III (p = 0.004) patients. Furthermore, multivariate analyses revealed that STYK1 protein expression was an independent prognostic indicator for both stage II (hazard ratio [HR], 2.472; p = 0.001) and stage III (HR, 2.001; p = 0.004) patients. CONCLUSIONS: Our results suggest that increased STYK1 protein expression correlates with disease progression and metastasis and may serve as a predictor of poor survival in CRC.

Upregulation of NETO2 expression correlates with tumor progression and poor prognosis in colorectal carcinoma.

BACKGROUND: Neuropilin and tolloid-like 2 (NETO2) has been found to be overexpressed in different human cancers, but its expression pattern and clinical relevance in colorectal carcinoma (CRC) remains unknown. METHODS: Real-time quantitative PCR, western blot and immunohistochemistry analyses were used to analyze the expression of NETO2 in CRC clinical samples. The correlation of NETO2 expression with clinicopathologic features was estimated in a cohort containing 292 patients with primary CRC. Kaplan-Meier and Cox proportional hazards regression analyses were used to assess the prognostic value of NETO2 expression in CRC. RESULTS: The expression of NETO2 was frequently upregulated in CRC clinical samples at both the mRNA and protein levels, and its upregulation was significantly correlated with poor tumor differentiation (p = 0.013), advanced local invasion (p = 0.049), increased lymph node metastasis (p = 0.009), advanced TNM stage (p = 0.041) and increased patient death (p = 0.001). Kaplan-Meier analysis of the complete study cohort revealed that patients with high-NETO2 tumors had a significantly shorter disease-specific survival (DSS) than those with low-NETO2 tumors (p < 0.001). Importantly, high levels of NETO2 protein predicted poor DSS for patients with early stage tumors (p = 0.027) and for those with advanced stage tumors (p = 0.020). Furthermore, multivariate analyses indicated that increased NETO2 expression was an independent unfavorable prognostic factor for patients with early stage tumors (hazard ratio [HR] = 1.937, 95% CI = 1.107-3.390, p = 0.021) as well as patients with advanced stage tumors (HR = 2.241, 95% CI = 1.245-4.035, p = 0.007). CONCLUSIONS: Our findings suggest that NETO2 upregulation could serve as a potential biomarker for the prediction of advanced tumor progression and unfavorable prognosis in patients with CRC.

Prognostic value of carbonic anhydrase VII expression in colorectal carcinoma.

BACKGROUND: Carbonic anhydrases (CAs) have been implicated in the pathogenesis of human cancers. Carbonic anhydrase VII (CA7), a member of the CA gene family, was recently demonstrated to be expressed in several human tissues including colon. Nevertheless, the expression and clinical relevance of CA7 in colorectal carcinoma (CRC) has not been investigated. METHODS: Real-time PCR, western blot, and immunohistochemistry analyses were used to determine CA7 expression in CRC clinical samples. The correlation of CA7 expression with clinicopathologic features was assessed in 228 patients from Luoyang, China (training cohort) and validated in 151 patients from Shanghai, China (validation cohort). Kaplan-Meier and Cox proportional regression analyses were used to estimate the association between CA7 expression and patients' survival. RESULTS: CA7 expression was frequently downregulated in CRC tissues at both the mRNA and protein levels. Reduced expression of CA7 was significantly correlated with poor differentiation, positive lymph node metastasis, advanced TNM stage and unfavorable clinical outcome not only in the training cohort but also in the validation set. Survival analysis indicated that patients with lower CA7 expression had a significantly shorter disease-specific survival (DSS) than those with higher CA7 expression. Importantly, further stage-based analyses revealed that decreased CA7 expression significantly predicted poor DSS and was an independent adverse prognostic indicator for patients with early stage tumors in both cohorts. CONCLUSIONS: Our results indicate that decreased expression of CA7 correlates with disease progression and predicts poor prognosis in CRC, especially for patients with early stage tumors.

Smad7 regulates compensatory hepatocyte proliferation in damaged mouse liver and positively relates to better clinical outcome in human hepatocellular carcinoma.

Transforming growth factor ß (TGF-ß) is cytostatic towards damage-induced compensatory hepatocyte proliferation. This function is frequently lost during hepatocarcinogenesis, thereby switching the TGF-ß role from tumour suppressor to tumour promoter. In the present study, we investigate Smad7 overexpression as a pathophysiological mechanism for cytostatic TGF-ß inhibition in liver damage and hepatocellular carcinoma (HCC). Transgenic hepatocyte-specific Smad7 overexpression in damaged liver of fumarylacetoacetate hydrolase (FAH)-deficient mice increased compensatory proliferation of hepatocytes. Similarly, modulation of Smad7 expression changed the sensitivity of Huh7, FLC-4, HLE and HLF HCC cell lines for cytostatic TGF-ß effects. In our cohort of 140 HCC patients, Smad7 transcripts were elevated in 41.4% of HCC samples as compared with adjacent tissue, with significant positive correlation to tumour size, whereas low Smad7 expression levels were significantly associated with worse clinical outcome. Univariate and multivariate analyses indicate Smad7 levels as an independent predictor for overall (P<0.001) and disease-free survival (P=0.0123). Delineating a mechanism for Smad7 transcriptional regulation in HCC, we identified cold-shock Y-box protein-1 (YB-1), a multifunctional transcription factor. YB-1 RNAi reduced TGF-ß-induced and endogenous Smad7 expression in Huh7 and FLC-4 cells respectively. YB-1 and Smad7 mRNA expression levels correlated positively (P<0.0001). Furthermore, nuclear co-localization of Smad7 and YB-1 proteins was present in cancer cells of those patients. In summary, the present study provides a YB-1/Smad7-mediated mechanism that interferes with anti-proliferative/tumour-suppressive TGF-ß actions in a subgroup of HCC cells that may facilitate aspects of tumour progression.

Crop plant genotyping by real-time PCR analysis of crude extracts of seeds.

Development of agricultural biotechnology requires rapid and convenient methods for crop plant genotyping. Real-time PCR is sensitive and reliable, and has been a routine technique in plant research. However, its application is limited by the cumbersome DNA template preparation procedures. We tested three PCR master mixes for direct amplification of crude seed DNA extracts without extensive purification. One mix had higher resistance to plant-derived PCR inhibitors and was shown to be applicable to various important crop plants. Furthermore, this method is capable of detecting single-copy genes from 2 mg pieces of seeds repetitively. Meanwhile, melting curve analysis could detect amplicons directly without electrophoresis manipulations. Taken together, this direct real-time PCR method provides a rapid and convenient tool for seed genotypic screening in crop plants.

Nuclear factor of activated T cells 5 maintained by Hotair suppression of miR-568 upregulates S100 calcium binding protein A4 to promote breast cancer metastasis.

INTRODUCTION: The onset of distal metastasis, which underlies the high mortality of breast cancers, warrants substantial studies to depict its molecular basis. Nuclear factor of activated T cells 5 (NFAT5) is upregulated in various malignancies and is critically involved in migration and invasion of neoplastic cells. Nevertheless, the metastasis-related events potentiated by this transcriptional factor and the mechanism responsible for NFAT5 elevation in carcinoma cells remain to be fully elucidated. METHODS: The correlation of NFAT5 with breast cancer invasiveness was investigated in vitro and clinically. The genes transcriptionally activated by NFAT5 were probed and their roles in breast cancer progression were dissected. The upstream regulators of NFAT5 were studied with particular attempt to explore the involvement of non-coding RNAs, and the mechanism underlying the maintenance of NFAT5 expression was deciphered. RESULTS: In metastatic breast cancers, NFAT5 promotes epithelial-mesenchymal transition (EMT) and invasion of cells by switching on the expression of the calcium binding protein S100A4, and facilitates the angiogenesis of breast epithelial cells and thus the development of metastases by transcriptionally activating vascular endothelial growth factor C (VEGF-C). NFAT5 is directly targeted by miR-568, which is in turn suppressed by the long non-coding RNA, Hotair, via a documented in trans gene silencing pattern, that is recruitment of the polycomb complex (Polycomb Repressive Complex 2; PRC2) and LSD1, and consequently methylation of histone H3K27 and demethylation of H3K4 on the miR-568 loci. CONCLUSION: This study unravels a detailed role of NFAT5 in mediating metastatic signaling, and provides broad insights into the involvement of Hotair, in particular, by transcriptionally regulating the expression of microRNA(s), in the metastasis of breast cancers.

Function characterization of a glyco-engineered anti-EGFR monoclonal antibody cetuximab in vitro.

AIM: To evaluate the biochemical features and activities of a glyco-engineered form of the anti-human epidermal growth factor receptor monoclonal antibody (EGFR mAb) cetuximab in vitro. METHODS: The genes encoding the Chinese hamster bisecting glycosylation enzyme (GnTIII) and anti-human EGFR mAb were cloned and coexpressed in CHO DG44 cells. The bisecting-glycosylated recombinant EGFR mAb (bisec-EGFR mAb) produced by these cells was characterized with regard to its glycan profile, antiproliferative activity, Fc receptor binding affinity and cell lysis capability. The content of galactose-α-1,3-galactose (α-Gal) in the bisec-EGFR mAb was measured using HPAEC-PAD. RESULTS: The bisec-EGFR mAb had a higher content of bisecting N-acetylglucosamine residues. Compared to the wild type EGFR mAb, the bisec-EGFR mAb exhibited 3-fold higher cell lysis capability in the antibody-dependent cellular cytotoxicity assay, and 1.36-fold higher antiproliferative activity against the human epidermoid carcinoma line A431. Furthermore, the bisec-EGFR mAb had a higher binding affinity for human FcγRIa and FcγRIIIa-158F than the wild type EGFR mAb. Moreover, α-Gal, which was responsible for cetuximab-induced hypersensitivity reactions, was not detected in the bisec-EGFR mAb. CONCLUSION: The glyco-engineered EGFR mAb with more bisecting modifications and lower α-Gal content than the approved therapeutic antibody Erbitux shows improved functionality in vitro, and requires in vivo validations.

Anesthetic isoflurane posttreatment attenuates experimental lung injury by inhibiting inflammation and apoptosis.

We investigated the effect of 1.4% isoflurane (ISO) on the development of inflammation and apoptosis caused by zymosan (ZY) in mice. We found that ZY-challenged mice exhibited significant body weight loss, markedly high mortality, and significant lung injury characterized by the deterioration of histopathology, histologic scores, and wet-to-dry ratio after ISO treatment. ISO dramatically attenuated ZY-induced lung neutrophil recruitment and inflammation, as evidenced by the reduced levels of total cells, neutrophils, and proinflammatory cytokines (i.e., tumor necrosis factor- α , interleukin- (IL-) 1 ß , IL-6, and macrophage inflammatory protein-2) in bronchoalveolar lavage fluid and of their mRNA expression in lung tissues. ISO also inhibited ZY-induced expression and activation of nuclear factor-kappaB p65 and inducible nitric oxide synthase in pulmonary tissue. ZY administration also resulted in the upregulation of heme oxygenase-1 expression and activity in the lung, which was further enhanced by ISO treatment. Moreover, ISO markedly prevented ZY-induced pulmonary cell apoptosis in mice, as reflected by the decrease in expression of procaspase-8, procaspase-3, cleaved caspase-8, and cleaved caspase-3, as well as in caspase-3 activity and Bcl-2-associated X/B-cell lymphoma 2 ratio. These results indicate that ISO is a potential therapeutic drug for treating ZY-induced lung injury, and further investigations are warranted.

Pulmonary fungal infection in a neonate with methylmalonic acidemia: A case report.

BACKGROUND: Methylmalonic acidemia (MMA) is characterized by non-specific symptoms such as vomiting, and feeding difficulties, along with delayed mental and physical development. However, no case of MMA combined with pulmonary fungal infection has been reported yet. CASE SUMMARY: We report the case of a neonate who presented pulmonary fungal infection along with the non-specific features of MMA. Exome sequencing revealed a c.331C>T variant in exon 3 of MMACHC from the father, and a c.658-c.660delAAG variant in exon 4 from the mother, which confirmed the diagnosis of cblC type MMA combined with hyperhomocysteinemia. CONCLUSION: Invasive fungal infection might occur in some infants with MMA. Therefore, early diagnosis is recommended for unexplained pulmonary infection.

Usability of Serum Stanniocalcin-1 as a Prognostic Biochemical Marker of Acute Supratentorial Intracerebral Hemorrhage: A Prospective Cohort Study.

Objective: Stanniocalcin-1 (STC1) may be neuroprotective. This study aimed to evaluate the prognostic role of serum STC1 levels in intracerebral hemorrhage (ICH). Methods: This prospective observational study was assigned in two parts. In the first part, blood samples of 48 patients with ICH were acquired on admission and on days 1, 2, 3, 5, and 7 after ICH, and those of 48 controls were collected at their entry into the study. In the second part, blood samples of 141 patients with ICH were obtained upon admission. Serum STC1 levels were measured, and the National Institutes of Health Stroke Scale (NIHSS), hematoma volume, and poststroke 6-month modified Rankin Scale (mRS) scores were recorded. Dynamic changes in serum STC levels and their correlation with disease severity and prognosis were investigated. Results: Serum STC1 levels were elevated after ICH, peaked on day 1, plateaued on day 2, declined gradually afterwards, and were significantly higher than those in controls. Serum STC1 levels were independently correlated with NIHSS scores, hematoma volume, and the 6-month post-injury mRS scores. Serum STC1 levels, NIHSS scores, and hematoma volume independently predicted a poor prognosis (mRS scores of 3-6). The model integrating serum STC1 levels, NIHSS scores, and hematoma volume was visually displayed using a nomogram and was relatively stable using the Hosmer-Lemeshow test and calibration curve analysis. Under the receiver operating characteristic curve, serum STC1 levels efficiently predicted a poor prognosis and showed similar prognostic ability to NIHSS scores and hematoma volume. The preceding model had significantly higher prognostic capability than NIHSS scores and hematoma volume alone and their combination. Conclusion: Substantial enhancement of serum STC1 levels after ICH, which is strongly correlated with severity, independently distinguished the risk of poor prognosis, assuming that serum STC1, as a prognostic parameter, may be clinically valuable in ICH.

Identification and assessment of new biomarkers for colorectal cancer with serum N-glycan profiling.

BACKGROUND: The objectives of this study were to identify and validate the diagnostic value of N-glycan markers in colorectal cancer (CRC) and to uncover their underlying molecular mechanism. METHODS: In total, 347 individuals, including patients with CRC, patients with colorectal adenoma, and healthy controls, were divided randomly into a training group (n = 287) and retrospective validation groups (n = 60). Serum N-glycan profiling was analyzed by DNA sequencer-assisted/flurophore-assisted carbohydrate electrophoresis (DSA-FACE). Two diagnostic models were constructed based on N-glycan profiling with logistic stepwise regression. The diagnostic performance of each model was assessed further in retrospective, prospective (n = 43), and follow-up (n = 46) cohorts. Lectin blot and reverse transcriptase-polymerase chain reaction were used to analyze the total core-fucosylated residues and molecular expression involved in core-fucosylation modifications in CRC. RESULTS: Two diagnostic models designated CRCglycoA and CRCglycoB were constructed to differentiate CRC from normal and adenoma, respectively. The areas under the receiver operating characteristic curves (AUC) of both CRCglycoA and CRCglycoB were higher than the AUC of carcinoembryonic antigen (CEA) (CRCglycoA, 0.92 vs 0.81; CRCglycoB, 0.81 vs 0.73). The sensitivity and accuracy of CRCglycoA improved from 21.7% to 25% and from 11.63% to 18% in the training cohort, the retrospective cohort, and the prospective cohorts compared with the sensitivity and accuracy of CEA. The sensitivity of CRCglycoB improved from 20% to 28.23%. Both altered N-glycans, and results from the diagnostic models were reversed after curative surgery. The level of total core fucose residues and fucosyltransferase were decreased significantly in CRC. CONCLUSIONS: The current results indicated that the N-glycan markers based diagnostic models are new, valuable, noninvasive alternatives for identifying CRC. The authors concluded that decreased fucosyltransferase may be responsible for decreased levels of total core-fucosylated modification in both tissues and serum from patients with CRC.

Long noncoding RNA high expression in hepatocellular carcinoma facilitates tumor growth through enhancer of zeste homolog 2 in humans.

UNLABELLED: In recent years, long noncoding RNAs (lncRNAs) have been shown to have critical regulatory roles in cancer biology. However, the contributions of lncRNAs to hepatitis B virus (HBV)-related hepatocellular carcinoma (HCC) remain largely unknown. Differentially expressed lncRNAs between HBV-related HCC and paired peritumoral tissues were identified by microarray and validated using quantitative real-time polymerase chain reaction. Liver samples from patients with HBV-related HCC were analyzed for levels of a specific differentially expressed lncRNA High Expression In HCC (termed lncRNA-HEIH); data were compared with survival data using the Kaplan-Meier method and compared between groups by the log-rank test. The effects of lncRNA-HEIH were assessed by silencing and overexpressing the lncRNA in vitro and in vivo. The expression level of lncRNA-HEIH in HBV-related HCC is significantly associated with recurrence and is an independent prognostic factor for survival. We also found that lncRNA-HEIH plays a key role in G(0) /G(1) arrest, and further demonstrated that lncRNA-HEIH was associated with enhancer of zeste homolog 2 (EZH2) and that this association was required for the repression of EZH2 target genes. CONCLUSIONS: Together, these results indicate that lncRNA-HEIH is an oncogenic lncRNA that promotes tumor progression and leads us to propose that lncRNAs may serve as key regulatory hubs in HCC progression.

Serum peptidome patterns of colorectal cancer based on magnetic bead separation and MALDI-TOF mass spectrometry analysis.

Background. Colorectal cancer (CRC) is one of the most common cancers in the world, identification of biomarkers for early detection of CRC represents a relevant target. The present study aims to determine serum peptidome patterns for CRC diagnosis. Methods. The present work focused on serum proteomic analysis of 32 health volunteers and 38 CRC by ClinProt Kit combined with mass spectrometry. This approach allowed the construction of a peptide patterns able to differentiate the studied populations. An independent group of serum (including 33 health volunteers, 34 CRC, 16 colorectal adenoma, 36 esophageal carcinoma, and 31 gastric carcinoma samples) was used to verify the diagnostic and differential diagnostic capability of the peptidome patterns blindly. An immunoassay method was used to determine serum CEA of CRC and controls. Results. A quick classifier algorithm was used to construct the peptidome patterns for identification of CRC from controls. Two of the identified peaks at m/z 741 and 7772 were used to construct peptidome patterns, achieving an accuracy close to 100% (>CEA, P < 0.05). Furthermore, the peptidome patterns could differentiate validation group with high accuracy. Conclusions. These results suggest that the ClinProt Kit combined with mass spectrometry yields significantly higher accuracy for the diagnosis and differential diagnosis of CRC.

Transforming growth factor beta1 gene variation Leu10Pro affects secretion and function in hepatic cells.

BACKGROUND: Our previous work revealed transforming growth factor beta1 (TGFß1) gene polymorphisms are associated with susceptibility to hepatocellular carcinoma and liver cirrhosis. However, no further study of functional substitution in hepatic cells has yet been reported. AIMS: This study was designed to uncover the functional mechanisms of TGFß1 gene polymorphisms in the pathogenesis of liver diseases. METHODS: Two recombinant TGFß1 expression plasmids containing TGFß1 codon 10 Leu/Pro variation were constructed with CMV promoter and transfected into human hepatoma cell lines (HepG2 and SMMU 7721), hepatic stellate cells (LX-2), and immortalized hepatocytes (L02). The secretion capacities of TGFß1 protein in the transfected cells were determined by ELISA. Apoptosis, proliferative activity, and expression of CD 105, CD83, and CD80 were also measured by use of flow cytometry. RESULTS: The ELISA results showed that cells transfected with CMV-Pro10 were more capable of TGFß1 secretion than those transfected with CMV-Leu10. Functionally, CMV-Pro10 was more apoptosis-protective and induced more proliferation than CMV-Leu10 in transfected hepatic cells. Pro10 up-regulated expression of CD105 and down-regulated expression of CD83. CONCLUSIONS: TGFß1 gene Leu10Pro variation in signal peptide has significant effects on TGFß1 secretion and functions in hepatic cells.

Serum peptidome patterns for early screening of esophageal squamous cell carcinoma.

Esophageal squamous cell carcinoma (ESCC) is one of the most common cancers in the world. Early diagnosis is critical for guiding the therapeutic management of ESCC. The present study aims to determine serum peptidome patterns for diagnosing ESCC. To identify novel peptidome patterns for diagnosing ESCC, sera from 31 healthy volunteers and 32 ESCC patients were subjected to a comparative proteomic analysis using a ClinProt™ Kit combined with mass spectrometry (MS). This approach enables the determination of peptidome patterns that can differentiate between ESCC sera and sera from healthy volunteers. For further validation, the diagnostic and differential diagnostic capabilities of the peptidome patterns were verified blindly by using an independent group of sera, consisting of sera from 31 ESCC patients, 33 healthy volunteers, 38 colorectal patients, and 36 gastric cancer patients. A Quick Classifier Algorithm was used to construct the peptidome patterns for the identification of ESCC from the control samples. Five of the identified peaks at mass to charge ratios 759, 786, 1,866, 3,316, and 6,634 were used to construct the peptidome patterns with almost 100% accuracy. Furthermore, the peptidome patterns could also differentiate the validation group with high accuracy. These results suggest that the ClinProt™ Kit combined with MS achieves significantly high accuracy for ESCC diagnosis and differential diagnosis.

Discovery and verification of gelsolin as a potential biomarker of colorectal adenocarcinoma in the Chinese population: Examining differential protein expression using an iTRAQ labelling-based proteomics approach.

OBJECTIVE: To identify and validate potential biomarkers of colorectal adenocarcinoma using a proteomic approach. METHODS: Multidimensional liquid chromatography/mass spectrometry was used to analyze biological samples labelled with isobaric mass tags for relative and absolute quantitation to identify differentially expressed proteins in human colorectal adenocarcinoma and paired normal mucosa for the discovery of cancerous biomarkers. Cancerous and noncancerous samples were compared using online and offline separation. Protein identification was performed using mass spectrometry. The downregulation of gelsolin protein in colorectal adenocarcinoma samples was confirmed by Western blot analysis and validated using immunohistochemistry. RESULTS: A total of 802 nonredundant proteins were identified in colorectal adenocarcinoma samples, 82 of which fell outside the expression range of 0.8 to 1.2, and were considered to be potential cancer-specific proteins. Immunohistochemistry revealed a complete absence of gelsolin expression in 86.89% of samples and a reduction of expression in 13.11% of samples, yielding a sensitivity of 86.89% and a specificity of 100% for distinguishing colorectal adenocarcinoma from normal tissue. CONCLUSIONS: These findings suggest that decreased expression of gelsolin is a potential biomarker of colorectal adenocarcinoma.