Simultaneous measurements of Cr, Cd, Hg and Pb species in ng L-1 levels by interfacing high performance liquid chromatography and inductively coupled plasma mass spectrometry.

Toxicity, mobility, bioavailability and biofunctions of chromium, cadmium, mercury and lead are heavily dependent upon their specific chemical forms, leading to a high demand to metal speciation analysis rather than total quantification. Simultaneous speciation analysis of multiple metal(loid)s is attractive to a large sample capacity containing unstable analytes due to its economic and environmental advantages over the conventional single elemental strategies. In this work, an analytical method integrating online solid phase extraction into high performance liquid chromatography interfaced with inductively coupled plasma mass spectrometry (ICP-MS) to simultaneously preconcentrate and quantify Cr, Cd, Hg and Pb forms in pg L-1 levels in water was developed. Cr(III + VI), Cd(II), Hg(II), Pb(II), methylmercury (MeHg), ethylmercury (EtHg), and trimethyl (TML) and triethyl lead (TEL) were captured by the C18 adsorbent (equilibrated with 10 mL of 1.0 mM 2-hydroxyethanethiol at 10 mL min-1) and eluted by mobile phase (5.0 mM Cys at pH 2.0), then completely separated on the C18 column within 8.0 min and eventually determined by ICP-MS. Low limits of detection (0.001-0.007 ng L-1) and quantification (0.003-0.023 ng L-1), good relative standard deviations (<4%) and high enrichment factors (827-2656 folds) were obtained with good linearities. Three reference materials of total cadmium (GBW08602), total mercury (GBW08603) and total lead (GBW08601) in water were analyzed by the developed method to validate the accuracy with good agreement with certified values and satisfactory recoveries (92-100%). This method was proved feasible by the determination of Cr, Cd, Hg and Pb compounds in drinking water, river water, pond water and tap water.

Simultaneous enrichment of inorganic and organic species of lead and mercury in pg L-1 levels by solid phase extraction online combined with high performance liquid chromatography and inductively coupled plasma mass spectrometry.

Quantification of ultra-trace inorganic and organic species of lead and mercury in unpolluted environmental water is crucial to estimate the mobility, toxicity and bioavailability and interactions. Simultaneous pre-concentration of Pb and Hg species in pg L-1 levels followed by multi-elemental speciation analysis makes great sense to a large set of unstable samples because of time advantages. Herein simultaneous enrichment and speciation analysis of ultra-trace lead and mercury in water was developed by online solid-phase extraction coupled with high performance liquid chromatography and inductively coupled plasma mass spectrometry (SPE-HPLC-ICP-MS) for this aim. Pb(II), trimethyl lead (TML), triethyl lead (TEL), Hg(II), methylmercury (MeHg) and ethylmercury (EtHg) were baseline separated in 11 min under gradient elution using 5 mM l-cysteine (Cys) at pH 2.5 in the 0-1 and 4-15 min and 5 mM Cys + 0.5 mM tetrabutyl ammonium hydroxide solution at pH 2.5 in the 1-4 min. Lead and mercury species in 10 mL intact water samples were adsorbed on a 1 cm C18 enrichment column pre-conditioned with 10 mL of 1 mM 2-mercaptoethanol at 10 mL min-1, and then directly desorbed by the mobile phases. High enrichment factors (459 for Pb(II), 1248 for TML, 1627 for TEL, 2485 for Hg(II), 1984 for MeHg and 1866 for EtHg) were obtained with good relative standard deviations (<5%), leading to low LODs (0.001-0.011 ng L-1) and LOQs (0.004-0.036 ng L-1). Good accuracy of this method was validated by two certified reference materials of total lead in water (GBW08601) and total mercury in water (GBW08603) along with spiked recoveries (89-93%). The method was applied to analyze trace lead and mercury species in river, lake, tap and rain water, and purified and mineral water. Inorganic lead of 13-68 ng L-1 and inorganic mercury of 21-49 ng L-1 were measured in the nine water samples whereas TML, TEL and MeHg were not detected with 2-5 ng L-1 EtHg presented only in one river water and tap water.

Quantification of ultra-trace organolead species in environmental water by inductively coupled plasma mass spectrometry with online solid-phase extraction and high performance liquid chromatographic separation.

Quantification of organolead compounds in environmental water is an essential task considering much higher toxicity and bioavailability of organolead species than inorganic plumbic ions. However, the speciation of ultra-trace organolead compounds at sub ng L-1 levels is challengeable for current instruments incorporating high performance liquid chromatography with inductively coupled plasma mass spectrometry (HPLC-ICP-MS) and even offline enrichment that offer detection limits around several to tens of ng L-1. In this paper, an online solid-phase extraction (SPE) coupled HPLC-ICP-MS method was developed for speciation analysis of trace lead in water. Graphene oxide bounded silica particles (GO@SiO2) was utilized as the SPE adsorbent because of its superior performance over graphene bounded silica particles and commercial C18 packing particles. High enrichment factors (1603 for TML and 1376 for TEL) were obtained when lead species in 10 mL sample was adsorbed by 1 mM sodium dodecyl benzene sulfonate (SDBS) preconditioned GO@SiO2 at 10 mL min-1 and then eluted by 5 µL of 5 mM SDBS. Because of the highly efficient preconcentration, detection limits were downscaled to be 0.018 for TML and 0.023 ng L-1 for TEL with relative standard deviations below 5%. Additionally, the proposed method also yielded rapid separation of Pb(II), TML and TEL (8 min) by using green mobile phases (aqueous solutions of 5 mM sodium 1-pentanesulfonate at pH 2.5 with/without 4 mM tetrabutylammounium hydroxide). Upon successful application to fresh water, TML and TEL were only presented in the river water whereas Pb(II) was only existed in the tap water, along with accuracy validation by good spiked recoveries (93-106%).

CircRNA_0005075 suppresses carcinogenesis via regulating miR-431/p53/epithelial-mesenchymal transition axis in gastric cancer.

This study was aimed to explore the expression and biological function of circRNA_0005075 in gastric cancer (GC) progression and its underlying mechanism. First, the expression level of circRNA_0005075 and microRNA-431 (miR-431) in GC tissues were detected with the quantitative real-time polymerase chain reaction. In addition, after down-regulated the circRNA_0005075 expression by plasmid transfection in GC cells, the Cell Counting Kit-8 (CCK-8), EDU, transwell assay were conducted to evaluate the function of circRNA_0005075 or miR-431 on cell proliferation, metastasis in vitro. Moreover, p53 and Epithelial-mesenchymal transition (EMT) pathway related proteins were also measured with western blotting. Then, our data revealed that CircRNA_0005075 was found to be significantly up-regulated in GC tissues as well as GC cell lines, and the GC patients with higher CircRNA_0005075 expression were more likely to have poor outcomes. Down-regulation of CircRNA_0005075 could significantly suppress the GC cell proliferation and cell metastasis ability, while the addition of miR-431 inhibitors could counteract this effect. Importantly, we discovered that the silencing of circRNA_0005075 could weaken the micro-RNA sponge function for miR-431, and then upregulate the expression of p53 and forbid the EMT signalling pathway, and finally suppress the tumourigenesis of GC. To sum up, CircRNA_0005075 could inhibit cell growth and metastasis of GC through regulating the miR-431/p53/EMT axis. SIGNIFICANCE OF THE STUDY: The research clearly elucidated the potential role and relative regulatory mechanism of circRNA_0005075 in gastric cancer (GC) progression. Briefly, circRNA_0005075 could directly inhibit the expression level of miR-431, then regulate the p53/Epithelial-mesenchymal transition axis, and finally inhibit cell growth and metastasis in GC. Consequently, circRNA_0005075 might act as an oncogene in the GC procession, which provides a promising way for the treatment of GC.

Traditional Chinese Medicine Curcumin Sensitizes Human Colon Cancer to Radiation by Altering the Expression of DNA Repair-related Genes.

BACKGROUND/AIM: The aim of the present study was to investigate the radio-sensitizing efficacy of curcumin, a traditional Chinese medicine (TCM) on colon cancer cells in vitro and in vivo. MATERIALS AND METHODS: Human colon cancer HT-29 cells were treated with curcumin (2.5 µM), irradiation (10 Gy) and the combination of irradiation and curcumin. Cell proliferation was assessed using the MTT assay. Apoptotic cells were detected by Annexin V-PE/7-AAD analysis. PCR was performed to determine differential-expression profiling of 95 DNA-repair genes in irradiated cells and cells treated with both irradiation and curcumin. Differentially-expressed genes were confirmed by Western blotting. In vivo radio-sensitizing efficacy of curcumin was assessed in a xenograft mouse model of HT-29 colon cancer. Curcumin was administrated daily by intraperitoneal injection at 20 mg/kg/dose. Mice received irradiation (10 Gy) twice weekly. Apoptosis of the cancer cells following treatment was determined by TUNEL staining. RESULTS: Irradiation induced proliferation inhibition and apoptosis of HT-29 cells in vitro. Concurrent curcumin treatment sensitized the HT-29 tumor to irradiation (p<0.01). DNA repair-related genes CCNH and XRCC5 were upregulated and LIG4 and PNKP downregulated by the combination of curcumin and irradiation compared with irradiation alone (p<0.05). Combined treatment of curcumin and irradiation resulted in a significantly greater tumor-growth inhibition and apoptosis compared to irradiation treatment alone (p<0.01). CONCLUSION: Curcumin sensitizes human colon cancer in vitro and in vivo to radiation. Downregulation of LIG4 and PNKP and upregulation of XRCC5 and CCNH DNA-repair-related genes were involved in the radio-sensitizing efficacy of curcumin in colon cancer.

[Study of the effect of LIG4 on the radiosensitivity enhancement of rectal cancer cells by curcumin].

OBJECTIVE: To investigate the function of repair gene LIG4 in radiosensitivity enhancement of rectal cancer cells by curcumin. METHODS: Human rectal cancer cells HT-29 were cultured in normal. LIG4-overexpression HT-29 cells and blank control plasmid HT-29 cells were established by gene transfection. Both kind of HF-29 cells were further randomly divided into curcumin group, radiotherapy group, curcumin plus radiotherapy group (combined group) and control group. The growth inhibition and apoptosis of cells were detected by MTT and Annexin V/PI respectively. Change of tumor volume was observed in nude mouse xenograft model, and the apoptosis of tumor cells was analyzed by TUNEL. RESULTS: Regarding blank control plasmid HT-29 cells, the growth inhibition rate and apoptosis rate in combined group were significantly higher than those in radiotherapy group(all P<0.05); tumor volume of nude mouse in combined group was significantly smaller than that in radiotherapy group, and the apoptotic index in combined group was significantly higher than that in radiotherapy group (all P<0.05). However, regarding LIG4-overexpression HT-29 cells, the growth inhibition rate and apoptosis rate were not significantly different between combined group and radiotherapy group(all P>0.05); the tumor volume of nude mouse and the apoptotic index were also not significantly different between combined group and radiotherapy group (all P>0.05). CONCLUSION: Down-regulation of LIG4 is an important mechanism of radiosensitivity enhancement of rectal cancer cells by curcumin.

[Mechanism of radiotherapy sensitization of curcumin on rectal cancer cells].

OBJECTIVE: To elucidate the mechanism of curcumin in radiotherapy sensitization for colorectal cancer cells. METHODS: Colorectal cancer HT-29 cells were cultured and treated with radiation and curcumin. MTT method was used to detect the cell growth inhibition. Then the high-throughput microarray was used to detect the differences in gene expression levels for each test group to identify differentially expressed genes, and each differential gene was validated by Western blotting. RESULTS: Cell growth inhibition rates at 48-hour and 72-hour in curcumin combined with radiotherapy group were significantly higher than those in simple radiotherapy group (P<0.05). Expression of 95 genes associated with gene-injury repair was detected by microarray. Compared to simple radiotherapy group, LIG4 and PNKP expression was down-regulated, and XRCC5 and CCNH expression was up-regulated in the curcumin combined with radiotherapy group (all P<0.05). Western blotting revealed LIG4 and PNKP protein expression decreased, and XRCC5 and CCNH protein expression increased in the curcumin combined with radiotherapy group as compared to the simple radiotherapy group (all P<0.05). CONCLUSION: Radiation sensitization effect of curcumin on colorectal cancer cells HT-29 may be associated with the regulation of genes of CCNH, LIG4, XRCC5, PNKP.

[Modified ligation of the intersphincteric fistula tract in the treatment of simple transsphincteric perianal fistula].

OBJECTIVE: To assess the efficacy and safety of modified ligation of the intersphincteric fistula tract for simple transsphincteric perianal fistula. METHODS: Seventy patients with simple transsphincteric perianal fistula between October 2012 and January 2014 in our department were prospectively enrolled. According to the random number table, patients were divided into two groups: modified-LIFT group (37 cases, from the external opening close to the fistula, dissect the external sphincter fistula to the intersphincteric groove by tunneling technique, resect the lateral free fistula) and LIFT group (33 cases). Clinical parametres before and after operation were compared, and results of pelvic electromyogram (EMG) and anorectal manometry three months after operation were analyzed to evaluated anal function. RESULTS: The operative time, pain score, hospital stay, and healing time were not significantly different between the two groups (all P>0.05). During the median follow-up of 12 months (3-20 months), the healing rate in modified-LIFT group was 83.8% (31/37), which was significantly higher than 60% (20/33) in LIFT group (P=0.029). After operation, 4 patients had persistent unhealed wound, 2 recurred in modified-LIFT group, while 8 patients had persistent unhealed wound, and 5 recurred in LIFT group. No patients developed anal incontinence. By the pelvic EMG and anorectal manometry 3 months after operation, the duration of motor unit potential, occurrence of simple phase, mean resting pressure and maximun squeeze pressure were not significantly different. CONCLUSION: Modified-LIFT procedure for the management of simple transsphincteric perianal fistulas is a simple and effective operation with higher healing rate and similar anal function as LIFT.

[Evaluation of anal function and quality of life after transanal endoscopic microsurgery].

OBJECTIVE: To evaluate the impact of transanal endoscopic microsurgery (TEM) on postoperative anal function and quality of life in patients with benign rectal tumor and early rectal cancer. METHODS: Clinical data of 50 patients with rectal adenoma and early rectal cancer undergoing transanal endoscopic microsurgery in our hospital from October 2008 to June 2013 were retrospectively analyzed. Anorectal manometry, endorectal ultrasonography (ERUS), the fecal incontinence severity index (FISI), and the physical and mental health status scores (SF-36) were used to evaluate preoperative and postoperative anorectal function and quality of life. RESULTS: Anorectal manometry indicated anal resting pressure (ARP), maximum squeeze pressure (MSP), rectal volume at sensory threshold(RVST), maximum tolerable volume(MTV) decreased significantly at the first month after surgery (P<0.05). MSP returned to preoperative level at the 3rd month (P>0.05). ARP and MTV returned to normal values at the 6th month (P>0.05). RVST returned to normal values at the 9th month (P>0.05). Recto-anal inhibitory reflex(RAIR) was absent in 1 (2%) patient preoperatively and in 30(60%), 18(36%), 7(14%), 2(4%) at the 1st, 3rd, 6th, 9th months after surgery respectively. ERUS showed similar width and thickness of internal sphincter at 1st and 6th month after surgery compared with preoperative measures (P>0.05). Six months after surgery, the mean FISI score decreased(preoperative vs postoperative:8.5 vs 5.8, P<0.05), suggesting an improvement in fecal continence. However, the overall quality of life did not danger significantly after surgery(P>0.05). CONCLUSIONS: TEM has little impact on anorectal anatomic structure. Anal function may be compromised in the short-term, however the vast majority of patients recover completely after 6-9 months. TEM is a safe, effective and minimally invasive surgery.