Co-culture of Acinetobacter sp. and Scedosporium sp. immobilized beads for optimized biosurfactant production and degradation of crude oil.

The widespread exploration and exploitation of crude oil has increased the prevalence of petroleum hydrocarbon pollution in the marine and coastal environment. Bioremediation of petroleum hydrocarbons using cell immobilization techniques is gaining increasing attention. In this study, the crude oil degradation performance of bacterial and fungal co-culture was optimized by entrapping both cells in sodium-alginate and polyvinyl alcohol composite beads. Results indicate that fungal cells remained active after entrapment and throughout the experiment, while bacterial cells were non-viable at the end of the experimental period in treatments with the bacterial-fungal ratio of 1:2. A remarkable decrease in surface tension from 72 mN/m to 36.51 mN/m was achieved in treatments with the bacterial-fungal ratio of 3:1. This resulted in a significant (P < 0.05) total petroleum hydrocarbon (TPH) removal rate of 89.4%, and the highest degradation of n-alkanes fractions (from 2129.01 mg/L to 118.53 mg/L), compared to the other treatments. Whereas PAHs removal was highest in treatments with the most fungal abundance (from 980.96 µg/L to 177.3 µg/L). Furthermore, enzymes analysis test revealed that catalase had the most effect on microbial degradation of the target substrate, while protease had no significant impact on the degradation process. High expression of almA and PAH-RHDa genes was achieved in the co-culture treatments, which correlated significantly (P < 0.05) with n-alkanes and PAHs removal, respectively. These results indicate that the application of immobilized bacterial and fungal cells in defined co-culture systems is an effective strategy for enhanced biodegradation of petroleum hydrocarbons in aqueous systems.

Enhanced remediation of oil-contaminated intertidal sediment by bacterial consortium of petroleum degraders and biosurfactant producers.

Oil pollution in intertidal zones is an important environmental issue that has serious adverse effects on coastal ecosystems. This study investigated the efficacy of a bacterial consortium constructed from petroleum degraders and biosurfactant producers in the bioremediation of oil-polluted sediment. Inoculation of the constructed consortium significantly enhanced the removal of C8-C40n-alkanes (80.2 ± 2.8% removal efficiency) and aromatic compounds (34.4 ± 10.8% removal efficiency) within 10 weeks. The consortium played dual functions of petroleum degradation and biosurfactant production, greatly improving microbial growth and metabolic activities. Real-time quantitative polymerase chain reaction (PCR) showed that the consortium markedly increased the proportions of indigenous alkane-degrading populations (up to 3.88-times higher than that of the control treatment). Microbial community analysis demonstrated that the exogenous consortium activated the degradation functions of indigenous microflora and promoted synergistic cooperation among microorganisms. Our findings indicated that supplementation of a bacterial consortium of petroleum degraders and biosurfactant producers is a promising bioremediation strategy for oil-polluted sediments.

Influence of microplastics on microbial anaerobic detoxification of chlorophenols.

Microplastics (MPs) can absorb halogenated organic compounds and transport them into marine anaerobic zones. Microbial reductive dehalogenation is a major process that naturally attenuates organohalide pollutants in anaerobic environments. Here, we aimed to determine the mechanisms through which MPs affect the microbe-mediated marine halogen cycle by incubating 2,4,6-trichlorophenol (TCP) dechlorinating cultures with various types of MPs. We found that TCP was dechlorinated to 4-chlorophenol in biotic control and polypropylene (PP) cultures, but essentially terminated at 2,4-dichlorophenol in polyethylene (PE) and polyethylene terephthalate (PET) cultures after incubation for 20 days. Oxygen-containing functional groups such as peroxide and aldehyde were enriched on PE and PET after incubation and corresponded to elevated levels of intracellular reactive oxygen species (ROS) in the microorganisms. Adding PE or PET to the cultures exerted limited effects on hydrogenase and ATPase activities, but delayed the expression of the gene encoding reductive dehalogenase (RDase). Considering the limited changes in the microbial composition of the enriched cultures, these findings suggested that microbial dechlorination is probably affected by MPs through the ROS-induced inhibition of RDase synthesis and/or activity. Overall, our findings showed that extensive MP pollution is unfavorable to environmental xenobiotic detoxification.

Enhanced bioremediation of diesel oil-contaminated seawater by a biochar-immobilized biosurfactant-producing bacteria Vibrio sp. LQ2 isolated from cold seep sediment.

This study investigated the effect of immobilized biosurfactant-producing bacteria on the bioremediation of diesel oil-contaminated seawater. Initially, a biosurfactant-producing bacterium, LQ2, was isolated from a marine cold-seep region, and identified as Vibrio sp. The biosurfactant produced by LQ2 was characterized as a phospholipid, exhibiting high surface activity with strong stability. Meanwhile, the inoculation of biochar-immobilized LQ2 demonstrated superior efficiency in removing diesel oil (94.7%, reduction from 169.2 mg to 8.91 mg) over a seven-day period compared to free-cell culture (54.4%), through both biodegradation and adsorption. In addition, the microbial growth and activity were greatly enhanced with the addition of immobilized LQ2. Further experiment showed that degradation-related genes, alkB and CYP450-1, were 3.8 and 15.2 times higher in the immobilized LQ2 treatment, respectively, than those in the free cell treatment. The findings obtained in this study suggest the feasibility of applying immobilized biosurfactant-producing bacteria, namely LQ2, in treating diesel oil-contaminated seawater.

An Analysis of Transcobalamin II Gene Polymorphisms and Serum Levels of Homocysteine, Folate and Vitamin B12 in Chinese Patients with Crohn's Disease.

OBJECTIVES: The study aimed to investigate the association of Crohn's disease (CD) with transcobalamin II (TCN2) polymorphisms and serum homocysteine, folate, and vitamin B12 levels. METHODS: TCN2 (rs1801198, rs9606756) were genotyped by iMLDR in 389 CD patients and 746 controls. Furthermore, 102 CD patients and 153 controls were randomly selected for examination of serum homocysteine, folate, and vitamin B12 levels by enzymatic cycling assay and chemiluminescence immunoassay, respectively. RESULTS: Mutant allele (G) and genotype (AG + GG) of (rs9606756) were higher in CD patients than in controls (both p < 0.05). So were they in ileocolonic CD patients and stricturing CD patients compared to controls (all p < 0.05). Mutant allele (G) and genotype (CG + GG) of (rs1801198) were more prevalent in stricturing CD patients than in controls (both p < 0.05). Compared to controls, average homocysteine level was enhanced in CD patients (p = 0.003), whereas average folate and vitamin B12 levels were reduced in CD patients (both p < 0.001). The prevalence of hyperhomocysteinemia, folate deficiency, and vitamin B12 deficiency was higher in CD patients than in controls (all p < 0.01). Both folate deficiency and vitamin B12 deficiency were independently related to risk of CD (both p < 0.01). CONCLUSION: TCN2 (rs1801198, rs9606756) polymorphisms as well as folate deficiency and vitamin B12 deficiency are correlated with CD.

Matrine inhibits BCR/ABL mediated ERK/MAPK pathway in human leukemia cells.

The BCR/ABL fusion gene and its downstream signaling pathways such as Ras/Raf/MAPK, JAK/STAT3, and PI3K/AKT pathways play important roles in malignant transformation of leukemia, especially chronic myelogenous leukemia (CML). Our previous study showed that matrine, an alkaloid extracted from a Chinese herb radix sophorae, significantly inhibited the proliferation of human CML K562cells, induced cell cycle arrest in G0/G1, and promoted cell apoptosis. In the present study, we investigated the molecular mechanism of matrine in the growth inhibition of leukemia cells using K562 and HL-60 cell lines. RT-PCR and Western blot assay demonstrated that the expression of BCR/ABL in K562 and HL-60 cells was significantly inhibited by matrine treatment. Phosphorylation of MEK1, ERK1/2, and their upstream adaptor molecules Shc and SHP2 were significantly downregulated. The protein and mRNA expression of components of the ERK/MAPK signal pathway, and Bcl-xL, Cyclin D1, and c-Myc, were dramatically reduced. Conversely, the expression of p27, a negative regulator of cell cycle progression, increased after matrine treatment. These results indicated that the inhibition of ERK/MAPK and BCR/ABL signaling pathway was associated with matrine's suppressive effects on the growth of K562 and HL-60 cells. In in vivo study, matrine significantly decreased the mortality rate of tumor-baring mice and suggested that matrine could exert its anti-leukemia effect in vivo.

Association of vitamin D receptor gene polymorphisms and serum 25-hydroxyvitamin D levels with Crohn's disease in Chinese patients.

BACKGROUND AND AIM: The vitamin D receptor (VDR) regulates immune responses and inflammation through binding with 1,25-dihydroxyvitamin D, the active form of vitamin D. The serum 25-hydroxyvitamin D (25(OH)D) level clinically reflects vitamin D status in the human body. We investigated the association of VDR polymorphisms and 25(OH)D levels in Chinese patients with Crohn's disease (CD). METHODS: Vitamin D receptor polymorphisms (FokI, BsmI, ApaI, and TaqI) were genotyped by SNaPshot. Serum 25(OH)D levels were measured by electro-chemiluminescence immunoassay. RESULTS: A total of 297 patients with CD and 446 controls were recruited. Compared with controls, mutant alleles and genotypes of BsmI and TaqI were less prevalent in patients with CD (all P < 0.05/4 = 0.0125). The AAC haplotype formed by BsmI, ApaI, and TaqI was also less prevalent in patients with CD (P = 0.004). Furthermore, 124 patients and 188 controls were randomly selected for measurements of 25(OH)D levels. Average 25(OH)D level was lower in patients with CD than in controls (15.46 ± 8.11 vs 21.64 ± 9.45 ng/mL, P < 0.001) and negatively linked to CD activity index (ß = -0.829, P < 0.001), platelet count (ß = -0.253, P < 0.001) and neutrophil percentage (ß = -0.136, P = 0.005) in patients with CD. The ApaI mutant genotype and vitamin D deficiency (<20 ng/mL) were independently associated with CD (P = 0.009, P < 0.001, respectively). In patients with CD, vitamin D deficiency interacted with FokI, ApaI, and TaqI mutant genotypes (P = 0.027, P = 0.024, and P = 0.040, respectively). CONCLUSIONS: Vitamin D receptor (BsmI, ApaI, and TaqI) mutations and lower 25(OH)D levels are associated with CD in Chinese patients. Moreover, VDR (FokI, ApaI, and TaqI) mutations and vitamin D deficiency may have a combined impact on CD.

STAT3 signaling pathway is involved in decitabine induced biological phenotype regulation of acute myeloid leukemia cells.

OBJECTIVE: This study aimed to investigate the role of signal transduction and transcriptional activator STAT3 and relevant signaling pathway in the DAC regulated biological phenotype of AML cells. METHODS: The effect of DAC at different concentrations on the proliferation of HL-60 cells was determined. After DAC treatment for 48 h, the killing capability of NK cells against HL-60 cells and the protein expressions of STAT3, JAK1, JAK2, SOCS-1 and SOCS-3 were evaluated. RESULTS: DAC markedly inhibited the proliferation of HL-60 cells. After the treatment of 48 hr with 0.2, 0.5 and 1.0 mol/L DAC, the HL-60 viability was reduced by 25±13%, 39±8% and 50±7% (P<0.01), respectively, and the early apoptosis rate was increased to 24.77±7.5%, 27.1±4.48% and 30.53±3.93%, respectively (control: 3.11±0.12%, P<0.01). DAC up-regulated the expression of MICA/B, ULBP-1 and ULBP-3 in HL-60 cells, and increased the killing activity of NK cells to HL-60 cells. DAC significantly induced the apoptosis of HL-60 cells and up-regulated the expression of NKG2D ligands in a dose dependent manner. Western blot assay showed the protein expression of STAT3, JAK, JAK2, phosphorylated STAT3, phosphorylated JAK1 and phosphorylated JAK2 decreased, while that of SOCS-1 and SOCS-3 increased in HL-60 cells after DAC treatment. CONCLUSION: In HL-60 cells, DAC can markedly inhibit their proliferation and up-regulate the expression of NKG2D ligands, and DAC also increase the cytotoxicity of NK cells to HL-60 cells, which may be related to the STAT3 related signaling pathway.

Matrine increases NKG2D ligand ULBP2 in K562 cells via inhibiting JAK/STAT3 pathway: a potential mechanism underlying the immunotherapy of matrine in leukemia.

PURPOSE: The study aimed to investigate the role of the JAK/STAT3 pathway in the matrine induced ULBP2 expression on the human chronic myelogenous leukemia K562 cells. METHODS: K562 cells were cultured, and the relevant mRNA expressions were detected. RESULTS: Matrine induced the expression of four NKG2D ligands on K562 cells, of which ULBP2 had the highest increase. After treatment with 0.8 mg/mL matrine for 24 h, the mean fluorescence intensity (MFI) of ULBP2 increased. After matrine treatment, the sensitivity of K562 cells to NK cell-mediated killing increased significantly. After treatment with 0.2, 0.5 and 0.8 mg/ mL matrine, the percentage of K562 cells killed by NK cells was significantly higher than that of untreated cells (29.2%) (P<0.05). Matrine significantly inhibit the protein expression of phosphorylated STAT 3 and JAK2. Matrine markedly inhibited the IL-6 expression of K562 cells, and antagonized the IL-6 mediated STAT3 and JAK2 phosphorylation. In addition, matrine enhanced the inhibitory effect of STAT 3 inhibitor on STAT 3 activity. The silencing of STAT expression and inhibition of STAT3 activity significantly up-regulated the ULPB2 expression. Matrine had no effect on the expression of IL-6R and gp130 on K562 cells, the mRNA expression of IL-6R and gp130 increased slightly and the sgp 130 in cell supernatant significantly increased. CONCLUSIONS: Our findings reveal IL-6 and IL-6 receptor-mediated JAK/STAT3 pathway is involved in the matrine induced up-regulation of NKG2D ligands ULBP2 on K562 cells. Matrine might inhibit IL-6 expression and then suppress the activation of IL-6 receptor-mediated JAK/STAT3 pathway.

[Association of inflammatory bowel disease with the polymorphisms and haplotypes of fucosyltransferase 3 gene].

OBJECTIVE: To assess the association of inflammatory bowel disease with polymorphisms and haplotypes of Fucosyltransferase 3 (FUT3) gene. METHODS: A total of 389 patients with ulcerative colitis (UC), 274 patients with Crohn's disease (CD), and 492 controls were collected. Three single nucleotide polymorphisms (SNPs) of the FUT3 gene (rs28362459, rs3745635 and rs3894326) were determined by direct sequencing. Linkage disequilibrium and haplotype analysis were performed using a Haploview 4.2 software. RESULTS: Compared with the controls, the allele and genotype distributions of FUT3 gene did not significantly differ between the UC and CD groups (all P>0.05). By stratified analysis, the mutant allele (A) and genotype (GA+AA) of the FUT3 gene (rs3745635) were significantly increased in the UC group with distal colitis compared with the controls (P<0.01, P<0.05, respectively). The mutant allele (G) and genotype (TG+GG) of the FUT3 gene (rs28362459) as well as the mutant allele (A) of FUT3(rs3745635) were significantly increased in patients with ileocolonic CD and ileal CD as compared with the controls (P<0.05, P<0.01, P<0.05, respectively). The frequency of mutant allele (G) of FUT3(rs28362459) was higher in stricturing CD patients than in the controls (P<0.05). In addition, the three polymorphic loci of FUT3 gene were shown in complete linkage disequilibrium [rs3894326/rs3745635 (D'=1.0, r2=0.017), rs3894326/rs28362459 (D'=0.937, r2=0.311), rs3745635/rs28362459 (D'=0.944, r2=0.448)]. However, the frequency of each haplotype was not significantly different between the UC and CD groups compared with the controls (all P>0.05). CONCLUSION: FUT3 (rs3745635) mutation may increase the risk of distal colitis. FUT3 (rs28362459 and rs3745635) mutations may engender the increased risk of ileocolonic and ileal CD. Moreover, FUT3 (rs28362459) polymorphism may influence the incidence of stricturing CD.

[An analysis of vitamin D receptor gene polymorphisms and serum 25-hydroxyvitamin D levels in patients with Crohn's disease].

OBJECTIVE: To investigate the association of Crohn's disease (CD) with vitamin D receptor (VDR) gene polymorphisms and serum 25-hydroxyvitamin D [25(OH)D] level. METHODS: A total of 297 CD patients and 446 healthy controls were enrolled in our study. Four single nucleosides of VDR (Fok I, Bsm I, Apa I and Taq I) were genotyped by SNaPshot. Serum 25(OH)D levels were tested by electro-chemiluminescence immunoassay in 124 CD patients and 188 matched random controls. RESULTS: By Chi-square test and Bonferroni correction, the frequencies of mutant allele (A) and mutant genotype (GA+AA) of Bsm I were significantly decreased in CD patients compared to controls [3.70% (22/594) vs 7.51% (67/892), 95% CI 0.289-0.776, P=0.002; 7.41%(22/297) vs 14.80% (66/446), 95% CI 0.277-0.765, P=0.002, respectively]. The similar results were seen for the mutant allele (C) and mutant genotype (TC+CC) of Taq I [4.21% (25/594) vs 7.62% (68/892), 95% CI 0.333-0.852, P=0.008; 8.42% (25/297) vs 14.57% (65/446), 95% CI 0.331-0.877, P=0.012]. The analyses of linkage disequilibrium (LD) and haplotype were performed by Haploview 4.2 and R software, respectively. The Bsm I, Apa I and Taq I polymorphic loci were found to be in a strong LD, and the AAC haplotype was significantly reduced in CD patients compared to controls [3.14% vs 6.46%, 95% CI 0.273-0.815, P=0.004]. The further serological analysis showed that average serum 25(OH)D level in CD patients was significantly lower than that of controls [(15.46±8.11) µg/L vs (21.64±9.45) µg/L, P<0.001]. By linear regression analysis, serum 25(OH)D levels in CD patients were negatively correlated to Crohn's disease activity index (ß=-0.829, P<0.001), platelet count (ß=-0.253, P<0.001) and the ratio of neutrophils (ß=-0.136, P=0.005) independently, whereas positively related to erythrocyte sedimentation rate (ß=0.191, P=0.001). Furthermore, logistic regression analysis was applied for establishing the models of gene-environment interaction. In result, both the mutant genotype (CA+AA) of Apa I and vitamin D deficiency (<20 µg/L) were shown to be the independent risk factors for CD (OR=7.580, 95% CI 2.983-19.261, P<0.001; OR=2.842, 95% CI 1.300-6.211, P=0.009, respectively). Besides, vitamin D deficiency in CD patients had multiplicative interactions with the mutant genotype (TC+CC) of Fok I, genotype (CA+AA) of Apa I and genotype (TC+CC) of Taq I, respectively (OR=0.419, 95% CI 0.194-0.906, P=0.027; OR=0.309, 95% CI 0.111-0.855, P=0.024; OR=5.841, 95% CI 1.082-31.538, P=0.040; respectively). CONCLUSIONS: VDR (Bsm I, Apa I and Taq I) polymorphisms and serum 25(OH)D levels are significantly related to CD. Both the mutant genotype (CA+AA) of Apa I and vitamin D deficiency are independent risk factors of CD. The mutations of VDR (Fok I, Apa I and Taq I) and vitamin D deficiency might have a synergistic effect on CD susceptibility.

[Associations of Crohn's disease with DR4 and DR5 gene polymorphisms].

OBJECTIVE: To assess the associations of death receptor DR4 and DR5 gene polymorphisms with Crohn's disease (CD). METHODS: A total of 295 CD patients and 490 healthy controls were recruited. Three single nucleotide polymorphisms (SNPs) of the DR4 (rs13278062, rs20575) and DR5 (rs1047266) genes were determined with a SNaPshot method. Unconditional logistic regression analysis was carried out for determining the allelic and genotypic differences of the three SNPs between CD patients and the controls, as well as the influence of the DR4 and DR5 gene polymorphisms on the clinical features of CD patients. Linkage disequilibrium and haplotype analysis were calculated by haplotype 4.2 and R language software. A gene-gene interaction model was established to analyze whether the three SNPs can exert a synergistic effect on the susceptibility to CD. RESULTS: The mutant allele (T) and genotype (GT+TT) of DR4 (rs13278062) were increased among CD patients compared to the controls (37.12% vs. 32.04%, P = 0.040, 95%CI: 1.010-1.550; 62.71% vs. 54.90%, P = 0.032, 95%CI: 1.028-1.855, respectively). However, the allelic and genotypic frequencies of DR4 (rs20575) and DR5 (rs1047266) did not differ between the two groups (all P > 0.05). Based on the Montreal Classification Standards, the CD patients were stratified by locations and behaviors of the disease. After multiple comparison correction (P < 0.0125), compared to ileocolonic CD patients respectively, the mutant allele (T) and genotype (GT+TT) of the rs13278062 polymorphism were significantly increased in colonic CD patients (41.04% vs. 25.64%, P = 0.002, 95%CI: 0.315-0.778; 66.04% vs. 41.03%, P = 0.001, 95%CI: 0.196-0.655, respectively) and terminal ileum CD patients (41.44% vs. 25.64%, P = 0.002, 95%CI: 0.311-0.762; 74.77% vs. 41.03%, P < 0.001, 95%CI: 0.126-0.437, respectively). In comparison to penetrating CD patients, the mutant allele (T) and genotype (GT+TT) of DR4 (rs13278062) were significantly decreased in stricturing CD patients (32.29% vs. 48.91%, P = 0.007, 95%CI: 0.300-0.828; 57.29% vs. 86.96%, P = 0.001, 95%CI: 0.078-0.520, respectively). A similar conclusion was drawn for the mutant genotype (GT+TT) of DR4 (rs13278062) in non-stricturing, non-penetrating CD patients (58.82% vs. 86.96%, P = 0.001, 95%CI: 0.086-0.536). Haplotype analysis indicated that the CT haplotype formed by rs20575 and rs13278062 was increased in CD patients compared to the controls (37.1% vs. 31.8%, P = 0.029, OR=1.279, 95%CI: 1.022-1.600). The outcome of a gene-gene interaction model indicated that the mutant genotype (GT+TT) of DR4 (rs13278062) and mutant genotype (CT+TT) of DR5 (rs1047266) may play a negatively synergistic role in CD patients (B = - 0.483, OR = 0.617, P = 0.030). CONCLUSION: The rs13278062 polymorphism of the DR4 gene not only can confer an increased risk for CD, but may also influence the location of the lesions and the disease behaviors. The CT haplotype formed by rs20575 and rs13278062 may be an independent risk factor for CD. Furthermore, the mutant genotype (GT+TT) of DR4 (rs13278062) and mutant genotype (CT+TT) of DR5 (rs1047266) may exert a negative synergistic effect on CD.

Matrine regulates immune functions to inhibit the proliferation of leukemic cells.

AIMS: To investigate the roles of matrine in regulating immune functions and its effect on the proliferation of leukemic cells. METHODS: Human leukemia K562, OUN-1, HL-60, U937, K562/AO2 cell lines and primary leukemic cells were used to detect the NKG2D ligands (NKG2DL) expression such as MICA/B, ULBP-1, ULBP-2, ULBP-3, and NK cells receptor NKG2D, CD158a, CD158b were detected by flow cytometry. Cell cytotoxic activity of human NK cells and CIK cells against K562 leukemia cells was detected using CFSE/PI double staining. Pro-inflammatory cytokines and adhesion molecules in K562 or NK cells supernatant after matrine treatment were detected. RESULTS: Matrine could upregulate the expression of NKG2DL on leukemic cell lines, and primary leukemic cells and enhance the NK and CIK cytotoxicity targeted to K562 cells. After matrine treatment, pro-inflammatory cytokines and adhesion molecular such as IL-6, IL-1, IL-2, IL-4, IL-5, GRO and TNF-α in K562 cells supernatant were significantly decreased (P < 0.05). Flow cytometry analysis showed that the NKG2D expression was up-regulated significantly as well as the CD158a and CD158b expression decreased after treatment with different concentration of matrine in a dose-dependent manner in K562 cells. A significant decrease of supernatant concentrations of IL-1α, IL-5, IL-6, IL-10, IFN-γ, GRO and TNF-α in NK cells was also observed after exposure to the matrine. CONCLUSION: Matrine regulates immune functions to inhibit the proliferation of leukemic cells.

[Growth inhibition effect of matrine on K562 cells mediated by IL-6/JAK/STAT3 signaling pathway].

OBJECTIVE: To investigate the molecular mechanism of the growth inhibitory effect of matrine on K562 cells in JAK/STAT3 mediated signal pathway. METHODS: Western blot analyses were performed to investigate the differential expression of JAK2, STAT3, phosphor-STAT3 (Tyr705 & Ser727) and phosphor-JAK2 proteins after matrine treatment in K562 cells with or without human recombinant interleukin 6 (IL-6) pretreatment. The expression of STAT3 response gene products such as Bcl-xL, Cyclin D1 and c-Myc, were investigated by Western blot and quantitative real time RT-PCR (qRT-PCR). Expression of IL-6, a potent upstream activating factor of JAK/STAT3 pathway, was analyzed by both real time qRT-PCR and ELISA. RESUTLS: Western blot revealed that matrine treatment resulted in a strong down-regulation of phosphor-STAT3 both in Tyr705 and Ser727 sites or phosphor-JAK2 proteins expression without significant effects on the total STAT3 and JAK2 proteins. The expression of phosphor-Tyr705 STAT3 and phosphor-Ser727 STAT3 was decreased to 0.370 ± 0.172 in K562 cells treated with 0.5 mg/ml matrine for 48 h, respectively, from 0.690 ± 0.119 and 1.150 ± 0.263 in control cells, accompanied with a dramatical down-regulation of phosphor-JAK2 from 0.670 ± 0.137 to 0.049 ± 0.057 (P<0.05). In addition, it was found that the expression of Bcl-xL, Cyclin D1, c-Myc was decreased both at the transcription and protein level in K562 cells after matrine treatment. Matrine treatment resulted in a significant decrease in the expression level of IL-6 in K562 cells from (35.1 ± 1.93) to (10.74 ± 1.83) and (8.66 ± 1.24) pg/ml at the dose of 0.5 and 0.8 mg/ml, respectively (p<0.05). Matrine treatment could diminish the up-regulation of STAT3, JAK2, phosphor-STAT3 and phosphor-JAK2 protein following pretreatment with IL-6 in K562 cells. CONCLUSION: Matrine exerts its anti-leukemia effect by interfering with the JAK2/STAT3 signaling pathway. The inhibition of IL-6 expression may play a pivotal role in the disruption of JAK/STAT pathway by matrine.

Matrine suppresses cell growth of human chronic myeloid leukemia cells via its inhibition of the interleukin-6/Janus activated kinase/signal transducer and activator of transcription 3 signaling cohort.

Matrine, alkaloid isolated from Sophora flavescens, is known to be pleiotropic by exerting anti-inflammatory, anti-oxidation, as well as anti-cancer effects. However, the precise molecular targets or pathways responsible for its activities still remain unclear. The present study aimed to determine the underlying mechanisms of matrine in inhibiting the chronic myeloid leukemia cells (CML). It was observed that matrine treatment significantly suppressed CML cells proliferation, induced apoptosis and resulted in the accumulation of cells in the G0/G1 phase, accompanied by a significant decrease in Bcl-xL, Cyclin D1, and c-Myc expression. Western blot analyses revealed that matrine treatment resulted in the down-regulation in phospho-STAT3 and phospho-JAK2 without significantly effects on STAT3 and JAK2 protein levels. Matrine significantly reduced the expression of IL-6, a potent upstream activating factor of STAT3. These results strongly suggested the IL-6/JAK/STAT3 pathway play an important role in matrine's anti-leukemia effects in K562 cells.

[Associations of the decoy receptor and osteoprotegerin gene polymorphisms with ulcerative colitis in Chinese patients].

OBJECTIVE: To investigate the correlation between decoy receptor (DcR)1, DcR2 and osteoprotegerin (OPG) gene polymorphisms with the susceptibility to ulcerative colitis (UC) in Chinese population. METHODS: A total of 352 patients with UC as well as 463 sex- and age-matched healthy controls were recruited in the study. The genetic polymorphisms of DcR1 (rs12549481), DcR2 (rs1133782) and OPG (rs3102735) were determined using a mini-sequencing technique method. RESULTS: In the autosomal dominant model, the rates of mutant allele (A) and genotype (GA+AA) of DcR2 (rs1133782) were lower in UC patients compared to the controls [6.25% (44/704) vs 8.96% (83/926), P = 0.043; 11.36% (40/352) vs 17.28% (80/463), P = 0.018, respectively]. In the recessive model, moreover, we found that the rates of mutant allele (T) and homozygote (TT) of OPG(rs3102735) were significantly increased in UC patients in contrast with the controls [86.36% (608/704) vs 81.53% (755/926), P = 0.009; 75.28% (265/352) vs 66.95% (310/463), P = 0.010, respectively]. By means of unconditional Logistic regression analysis, the rate of mutant allele (T) of OPG (rs3102735) was shown to be significantly decreased in patients with severe UC compared to the other UC patients [76.67% (69/90) vs 87.79% (539/614), OR = 0.457, 95%CI 0.265-0.788, P = 0.004]. Nevertheless, the genetic polymorphism of DcR2(rs1133782) was not significantly related to the clinical features in UC patients. In addition, the genotypic distribution of DcR1 (rs12549481) in control group did not conform to the Hardy-Weinberg equilibrium rule, thus a further statistical analysis was not performed in our study. CONCLUSIONS: The genetic polymorphism of DcR2(rs1133782) might be associated with the susceptibility to UC. Not only is the mutation of OPG (rs3102735) gene correlated to the development of UC, but also to the severity of disease.

Associations of FUT2 and FUT3 gene polymorphisms with Crohn's disease in Chinese patients.

BACKGROUND AND AIM: FUT2 and FUT3 genes are responsible for the formation of histo-blood group antigens, which act as binding sites for some intestinal microbes. Several studies suggested that FUT2 gene might affect the intestinal microbiota composition and modulate innate immune responses. However, the effect of FUT2 polymorphisms on Crohn's disease (CD) is uncertain. Our study aimed to analyze associations of CD with FUT2 and FUT3 polymorphisms in Chinese population. METHODS: A total of 273 CD patients and 479 controls were recruited. The genotypes of FUT2 (rs281377, rs1047781, and rs601338) and FUT3 (rs28362459, rs3745635, and rs3894326) were detected by SNaPshot analysis. RESULTS: Compared with controls, homozygote TT of FUT2 (rs1047781) was significantly increased in CD patients (TT vs others; P = 0.002, odds ratio [OR] = 1.767, 95% confidence interval [CI] = 1.235-2.528). The haplotype TT formed with FUT2 (rs281377) and (rs1047781) was more prevalent in CD patients than in controls (48.9% vs 43.5%, P = 0.046). Mutant T allele and homozygote TT of FUT2 (rs1047781) were increased in colonic CD patients compared with controls (P < 0.001, OR = 1.843, 95% CI = 1.353-2.512; P < 0.001, OR = 2.607, 95% CI = 1.622-4.191, respectively). Although allele and genotypic distributions of FUT3 were not statistically different between CD patients and controls, mutant allele and genotype of FUT3 (rs28362459) and (rs3745635) were significantly discrepant in three subgroups of CD patients according to lesion locations (all P < 0.05). CONCLUSIONS: Our study strongly implicates the polymorphic locus of FUT2 (rs1047781) in CD susceptibility in Chinese population. Mutations of FUT3 (rs28362459) and (rs3745635) might influence the lesion locations in CD patients.

Highly efficient and recyclable carbon soot sponge for oil cleanup.

Carbon soot (CS) has the advantages of cost-effectiveness and production scalability over other carbons (i.e., graphene, CNTs) in their synthesis. However, little research has been conducted to explore the potential applications of CS. In this study, we demonstrated that a common daily waste-CS-can be used for developing a cost-effective absorbent (CS-sponge) to remove oil contaminants from water. The CS was synthesized by an ethylene-oxygen combustion flame. The CS-sponge was prepared via a dip-coating method. Without further surface modification and pretreatments, the CS-sponge demonstrates high absorption capacities (up to 80 times its own weight) for a broad spectrum of oils and organic solvents with a recyclability of more than 10 times. These research results show evidence that the CS-sponge is promising in environmental remediation for large-scale, low-cost removal of oils from water.

[The associations of ulcerative colitis with vascular endothelial growth factor (-2578C/A) and (+936C/T) single nucleotide polymorphisms].

OBJECTIVE: To investigate the association of (-2578C/A) and (+936C/T) single nucleotide polymorphism(SNPs) of vascular endothelial growth factor (VEGF) gene with the susceptibility to ulcerative colitis (UC). METHODS: A total of 373 UC patients and 503 healthy controls were recruited. The (-2578C/A) and (+936C/T) polymorphism of VEGF gene were detected using a mini-sequencing technique. RESULTS: By an unconditional logistic regression analysis, the frequencies of the mutant allele T and genotype CT+TT of VEGF gene (+936C/T) were significantly decreased in patients with severe UC compared to the controls (10.4% vs 19.3%, OR = 0.487, 95%CI 0.248-0.954, P = 0.036; 18.8% vs 33.8%, OR = 0.452, 95%CI 0.214-0.955, P = 0.037, respectively). Moreover, patients with severe UC had significant lower rates of mutant allele T and genotype CT+TT compared with patients with mild and moderate UC (10.4% vs 20.5%, OR = 0.452, 95%CI 0.229-0.894, P = 0.022; 18.8% vs 36.9%, OR = 0.394, 95%CI 0.185-0.842, P = 0.016, respectively). The frequencies of mutant allele A and genotype CA+AA of VEGF (-2578C/A) gene were not statistically different between UC patients and the controls. Moreover, they were not significantly associated with the clinicopathologic features in UC patients. CONCLUSIONS: The mutation of VEGF (+936C/T) gene is correlated with the severity of UC. However, the polymorphism of VEGF (-2578C/A) gene is not significantly related to the susceptibility to UC.

[Effects of matrine on the expression of NKG2D ligands in leukemia cells].

This study was aimed to analyze the expression of NKG2D ligands in human leukemic cells and to investigate the effects of matrine on NKG2D ligand expression. The expressions of NKG2D ligand MICA/B, ULBP1-3 in several human leukemia cell lines (K562, OUN-1, U937 and K562/AO2), as well as primary leukemic cells isolated from malignant leukemia patients were analyzed by flow cytometry. After treatment with different doses of matrine, the expression level of NKG2D ligands in these leukemic cells was detected by FCM. The results indicated that NKG2D ligand expression was detected in both the leukemia cell lines and primary malignant leukemic cells. Generally, the expression of ULBP was high or obviously higher than that of MICA/B in leukemia cell lines and primary leukemic cells. The expression pattern of NKG2D ligands was different among these cells, possibly due to the different types of leukemia. Not all the expression of NKG2D ligands was upregulated after matrine treatment. Much higher expressions of ULBP2 and ULBP3 were found in K562 cells, compared to the other cell lines, which partly contributes to the higher sensitivity of K562 cells to NK cytotoxicity as target cells. It is concluded that there is universal expression of NKG2D ligand in leukemia cells. The high ULBP expression is prevalent in human leukemia cells. Matrine has the potential to induce the expression of NKG2D ligands in leukemia cells.