Efficacy and prognosis of adalimumab for Crohn's disease at different disease locations: a systematic review and meta-analysis.

OBJECTIVE: For over ten years, adalimumab (ADA) has been approved for treating active moderate to severe Crohn's disease (CD), showing irreplaceable efficacy. However, the difference in efficacy and prognosis when the disease pathology occurs in different locations of the body is still unclear. This study used systematic meta-analysis to assess the efficacy of ADA and prognosis in CD in different locations of disease pathology. MATERIALS AND METHODS: We used "Adalimumab OR ADA OR HUMIRA OR IgG1 monoclonal antibody" AND "Crohn disease OR Crohn's disease OR CD OR IBD OR inflammatory bowel disease" as search strategies for searching electronic databases in the Embase, PubMed and CNKI databases. A systematic meta-analysis of proportions was performed to analyze the data. RESULTS: A total of 1,253 patients in 15 articles were included in our study. The results showed that treatment with ADA led to overall remission rates that were elevated (70%, 95% CI: 58%-79%) but a nonnegligible overall relapse rate (28%, 95% CI: 12%-53%) in patients with CD. More importantly, we indicated that the use of ADA in patients with colon only (L2), ileum and colon (L3) and upper gastrointestinal tract (L4) CD led to significantly lower clinical remission rates. The use of ADA in patients with L2 led to significantly higher relapse rates, but the use of ADA in patients with ileum only (L1) and L3 CD led to significantly lower relapse rates. CONCLUSIONS: Our findings clarify different remission and relapse rates depending on the location of the disease pathology and may be useful for clinicians' choice of treatment strategies.

[Laparoscopic diagnosis of postoperative recurrence of peritoneal metastasis in gastric cancer patients and the clinical efficacy of bidirectional intraperitoneal and systemic chemotherapy].

Objective: To explore the diagnostic value of laparoscopy in the postoperative recurrence of peritoneal metastasis in gastric cancer, and to investigate the efficacy of bidirectional intraperitoneal and systemic (BIPS) chemotherapy for the recurrence. Methods: The descriptive case series study was conducted. Case inclusion criteria: (1) gastric cancer patients without synchronous distant metastasis received D2 radical gastrectomy; (2) postoperative adjuvant chemotherapy was administered; (3) no other distant metastasis except recurrence of peritoneal metastasis; (4) age of 18-75 years; (5) Eastern Cooperative Oncology Group (ECOG) performance-status score≤2; (6) pretreatment evaluation suggested that surgery and chemotherapy could be tolerated. Eight consecutive gastric cancer patients with postoperative recurrence of peritoneal metastasis who met the above criteria at Department of Gastrointestinal Surgery of Ruijin Hospital from September 2015 to September 2016 were enrolled into the study. There were 6 males and 2 females with the median age of 52 (38-68) years. They received laparoscopy or laparotomy first, and then were evaluated with reference to the Sugarbaker peritoneal cancer index (PCI) and the peritoneal metastasis classification of gastric cancer developed by the Japanese Gastric Cancer Research Association. A peritoneal access port was implanted in the subcutaneous space of the lower abdomen and the patients received chemotherapy for 21 days as a course of treatment. All the patients received intraperitoneal 20 mg/m(2) of paclitaxel (PTX) via implanted subcutaneous peritoneal access ports and intravenous 50 mg/m(2) of PTX at day 1 and day 8, meanwhile 80 mg/m(2) of Tigio was orally administered per day for 14 consecutive days, followed by 7 days of interval. Follow-up ended on December 15, 2019. Results: Of these 8 patients with recurrence of peritoneal metastasis after gastric cancer surgery, 1 case underwent laparotomy and loop stoma of terminal ileum because of complete colonic obstruction, and the remaining 7 cases underwent laparoscopy successfully and the recurrence of peritoneal metastasis was clearly diagnosed. Two patients with ovarian metastasis underwent laparoscopic bilateral adnexectomy. The median follow-up time was 17.5 (1.5 to 39.0) months, the median number of BIPS chemotherapy course was 11 (1 to 30), and the median survival time (MST) after BIPS chemotherapy was 17.0 months. The major adverse reaction in BIPS treatment was mainly myelosuppression, of which grade 3/4 leukopenia and neutropenia developed in 1 and 2 cases respectively. No BIPS-related death occurred. The MST of gastric cancer after radical gastrectomy was 40.0 months. Conclusions: Laparoscopy is a safe and feasible method for diagnosing the recurrence of peritoneal metastasis of gastric cancer. BIPS chemotherapy is effective and safe for its treatment and deserves further study.

Relative survival of Bacillus subtilis spores loaded on filtering facepiece respirators after five decontamination methods.

This study determines the relative survival (RS) of Bacillus subtilis spores loaded on an N95 filtering facepiece respirator (FFR) after decontamination by five methods under worst-case conditions. Relative survival was obtained by testing after decontamination and after storing the FFRs at 37°C and 95% relative humidity for 24 hours. The decontamination methods involved ethanol, bleach, ultraviolet irradiation (UVA 365 nm, UVC 254 nm), an autoclave, and a traditional electric rice cooker (TERC) that was made in Taiwan. Without decontamination, 59 ± 8% of the loaded spores survived for 24 hours. When 70% ethanol was added to the N95 FFR at a packing density of 0.23, the RS was 73 ± 5% initially and decayed to 22 ± 8% in 24 hours. Relative survival remained above 20% after 20 minutes of UVA irradiation. The other four decontamination measures achieved 99%-100% biocidal efficacy, as measured immediately after the methods were applied to the test FFRs. Relative survival is a useful parameter for measuring sterilization or degree of disinfection. Bleach, UVC, an autoclave, and a TERC provide better biocidal efficacy than ethanol and UVA. Not only a higher filter quality but also a lower value of RS produced the most decontaminated FFR.

Drug Target Identification Using an iTRAQ-Based Quantitative Chemical Proteomics Approach-Based on a Target Profiling Study of Andrographolide.

Identifying the cellular binding targets of drugs and other bioactive small molecules is a crucial step for understanding their molecular mechanisms of action as well as potential off-target effects. The field of chemical proteomics is an emerging discipline in chemical biology using synthetic chemistry and high-throughput detection techniques to study small molecule-protein interactions. In this chapter, we describe a quantitative chemical proteomics protocol combining bioorthogonal click chemistry and quantitation by isobaric tags for relative and absolute quantification (iTRAQ) to identify the specific binding targets of drugs and bioactive small molecules such as natural products. A modified drug probe with a click chemistry-enabling addition is synthesized and used in live cell treatments where it undergoes covalent interactions with its cognate cellular targets. The probes are then ligated to biotin through click chemistry and enriched with avidin beads, followed by iTRAQ labeling and liquid chromatography-tandem mass spectrometry (LC-MS/MS) analysis for protein identification and relative quantitation discriminating specific targets from nonspecific binding proteins. The presented protocol has been used to successfully profile prominent drugs and natural products including andrographolide, aspirin, curcumin, etc., and can be a powerful tool to study the molecular mechanisms of bioactive small molecules.

Mitochondrial inhibitor sensitizes non-small-cell lung carcinoma cells to TRAIL-induced apoptosis by reactive oxygen species and Bcl-X(L)/p53-mediated amplification mechanisms.

Tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) is a promising agent for anticancer therapy; however, non-small-cell lung carcinoma (NSCLC) cells are relatively TRAIL resistant. Identification of small molecules that can restore NSCLC susceptibility to TRAIL-induced apoptosis is meaningful. We found here that rotenone, as a mitochondrial respiration inhibitor, preferentially increased NSCLC cells sensitivity to TRAIL-mediated apoptosis at subtoxic concentrations, the mechanisms by which were accounted by the upregulation of death receptors and the downregulation of c-FLIP (cellular FLICE-like inhibitory protein). Further analysis revealed that death receptors expression by rotenone was regulated by p53, whereas c-FLIP downregulation was blocked by Bcl-X(L) overexpression. Rotenone triggered the mitochondria-derived reactive oxygen species (ROS) generation, which subsequently led to Bcl-X(L) downregulation and PUMA upregulation. As PUMA expression was regulated by p53, the PUMA, Bcl-X(L) and p53 in rotenone-treated cells form a positive feedback amplification loop to increase the apoptosis sensitivity. Mitochondria-derived ROS, however, promote the formation of this amplification loop. Collectively, we concluded that ROS generation, Bcl-X(L) and p53-mediated amplification mechanisms had an important role in the sensitization of NSCLC cells to TRAIL-mediated apoptosis by rotenone. The combined TRAIL and rotenone treatment may be appreciated as a useful approach for the therapy of NSCLC that warrants further investigation.

FADD regulates thymocyte development at the ß-selection checkpoint by modulating Notch signaling.

Non-apoptotic functions of Fas-associated protein with death domain (FADD) have been implicated in T lineage lymphocytes, but the nature of FADD-dependent non-apoptotic mechanism in early T-cell development has not been completely elucidated. In this study, we show that tissue-specific deletion of FADD in immature (CD44(-)CD25(+)) thymocytes results in severe perturbation of αß lineage development. Meanwhile, loss of FADD signaling at a later (CD44(-)CD25(-)) developmental stage does not affect subsequent T-cell development. Collectively, our work presents that FADD deficiency induces failed survival in double-negative 4 (DN4) cells, while pre-T-cell receptor (TCR) signal remains intact. In addition, Notch signaling is positive regulated on DN4 and double-positive thymocytes in T-cell-specific FADD-knockout mice, which express higher levels of a subset of Notch-target genes, including Hes1, Deltex1 and CD25. Moreover, a transcriptional repressor of Notch1, NKAP is downregulated coupled with the loss of FADD in thymocytes and is found to associate with FADD. These data suggest that as a death receptor, FADD is also required for cell survival in ß-selection as a regulator of Notch1 expression.

Suppression of tumor angiogenesis by targeting the protein neddylation pathway.

Inhibition of protein neddylation, particularly cullin neddylation, has emerged as a promising anticancer strategy, as evidenced by the antitumor activity in preclinical studies of the Nedd8-activating enzyme (NAE) inhibitor MLN4924. This small molecule can block the protein neddylation pathway and is now in clinical trials. We and others have previously shown that the antitumor activity of MLN4924 is mediated by its ability to induce apoptosis, autophagy and senescence in a cell context-dependent manner. However, whether MLN4924 has any effect on tumor angiogenesis remains unexplored. Here we report that MLN4924 inhibits angiogenesis in various in vitro and in vivo models, leading to the suppression of tumor growth and metastasis in highly malignant pancreatic cancer, indicating that blockage of angiogenesis is yet another mechanism contributing to its antitumor activity. At the molecular level, MLN4924 inhibits Cullin-RING E3 ligases (CRLs) by cullin deneddylation, causing accumulation of RhoA at an early stage to impair angiogenic activity of vascular endothelial cells and subsequently DNA damage response, cell cycle arrest and apoptosis due to accumulation of other tumor-suppressive substrates of CRLs. Furthermore, we showed that inactivation of CRLs, via small interfering RNA (siRNA) silencing of its essential subunit ROC1/RBX1, recapitulates the antiangiogenic effect of MLN4924. Taken together, our study demonstrates a previously unrecognized role of neddylation in the regulation of tumor angiogenesis using both pharmaceutical and genetic approaches, and provides proof of concept evidence for future development of neddylation inhibitors (such as MLN4924) as a novel class of antiangiogenic agents.

Using c-Fos/c-Jun as hetero-dimer interaction model to optimize donor to acceptor concentration ratio range for three-filter fluorescence resonance energy transfer (FRET) measurement.

Sensitized emission FRET detection method based on three-filter fluorescence microscopy is widely used and more suitable for live cell FRET imaging and dynamic protein-protein interaction analysis. But when it is applied to detect two proteins interaction in living cells, this intensity-based detection method is complicated by many experimental factors such as spectral crosstalk and spectral bleed-through and variable donor to acceptor concentration ratio. There are several FRET algorithms developed recently to correct those factors in order to quantitatively gauge and compare FRET signals between different experimental groups. But the algorithms are often difficult to choose when they are applied to certain experiments. In this research, we use c-Fos/c-Jun as a simple hetero-dimer interaction model to quantitatively detect and compare the FRET signals based on the following widely used sensitized emission FRET algorithms: N(FRET) , FRET(N) , FR, FRET(R) , E(app) and E(EFF) . We optimized the donor to acceptor concentration ratio range for the above FRET algorithms and facilitate their use in accurate FRET signal determination based on the three-filter FRET microscopy.

Peripheral blood dendritic cells and vascular endothelial growth factor in oral squamous cell carcinoma: correlation analysis and in vitro study.

Vascular endothelial growth factor (VEGF) may cause functional deficiency in dendritic cells (DCs) in vitro. The roles of peripheral blood dendritic cells (PBDCs) and VEGF in patients with oral squamous cell carcinoma (OSCC) are not well understood. The authors analysed the correlation between VEGF and PBDC in 81 OSCC patients. They assessed the effect of VEGF on DC function in vitro. VEGF levels were significantly increased in OSCC patients compared with control subjects (P < 0.05), but PBDC levels were significantly lower (P < 0.05). VEGF expression in TNM I-II (67%) and T1-T2 (74%) was significantly lower, compared with TNM III-IV (88%, P < 0.05) and T3-T4 (89%, P < 0.05). Increased VEGF expression in primary tumours was significantly correlated with elevated serum VEGF levels, but reduced PBDC levels. In vitro cultured DC exposed to VEGF showed significantly decreased expression of functional proteins, enhanced endocytosis activity, and elicited weaker proliferation of T cells, compared with that of free VEGF (P < 0.01). These findings suggest that decreased PBDC and elevated VEGF occur in OSCC patients. Higher VEGF levels may affect precursor cells, resulting in decreased numbers of functional DC.

Impairment of Na(+),K(+)-ATPase in CD95(APO-1)-induced human T-cell leukemia cell apoptosis mediated by glutathione depletion and generation of hydrogen peroxide.

Human T-cell leukemia is a malignant disease that needs various regimens of cytotoxic chemotherapy to overcome drug resistance. Recently, Na(+),K(+)-ATPase has emerged as a potential target for cancer therapy. However, its exact signaling pathway in human T-cell leukemia cell death has not been well defined. In the current study, we found CD95(APO-1) was able to trigger the internalization of plasma membrane Na(+),K(+)-ATPase in Jurkat cells or primary T cells as a mechanism to suppress its activity. This internalization was closely relevant to intracellular glutathione (GSH) depletion in Jurkat cells downstream of Fas-associated death domain protein (FADD) and caspase 8. GSH depletion in Fas L-treated Jurkat cells induced the generation of hydrogen peroxide (H(2)O(2)), which subsequently increased the serine phosphorylation of Na(+),K(+)-ATPase alpha1 subunit. Exogenous H(2)O(2) even mimicked the effect of Fas L to upregulate the serine phosphorylation of Na(+),K(+)-ATPase alpha1 subunit and suppress Na(+),K(+)-ATPase activity. Overall, our results indicate that CD95(APO-1) induces the FADD- and caspase 8-dependent internalization of Na(+),K(+)-ATPase through intracellular GSH loss, and the subsequent generation of H(2)O(2)-mediated serine phosphorylation of Na(+),K(+)-ATPase alpha1 subunit. Taken together, this study presents a novel regulatory mechanism of Na(+),K(+)-ATPase in CD95(APO-1)-mediated human T-leukemia cell apoptosis.

Extracts from Plastrum testudinis promote proliferation of rat bone-marrow-derived mesenchymal stem cells.

OBJECTIVES: The purpose of this study is to identify active components of PT involved in promoting proliferation of MSCs and to investigate its mechanism. PT was extracted with petroleum ether, ethyl acetate, ethanol and water respectively. MATERIALS AND METHODS: Evidence provided by MTT, HE stain, BrdUrd, PCNA immunoreactivity and cell cycle indicated that Plastrum Testudinis Extracted with ethyl acetate (PTE) is the only active components responsible for increasing MSCs proliferation. RESULTS: This finding leads us to identify the chemical component of PTE. Steroid, fatty acids and their esters components in PTE were determined by GC-MS and HPLC. The mechanism of PTE action may be associated with the up-regulation of BMP4. CONCLUSIONS: Our findings give novel insights into the promoting effects of Plastrum Testudinis on proliferation of MSCs and help to identify the chemical component and to clarify the mechanism of its pharmacological activities.

Selecting peptide ligands of microcystin-LR from phage displayed random libraries.

In the present study, we investigated to find novel ligands for low molecular weight environmental toxin, microcystin-LR (MC-LR) by using phage display technology. Two random libraries, displaying linear 12-mer peptides and cyclic 7-mer peptides, were screened against the immobilized target respectively. After three rounds of panning, phage clones that recognized microcystin-LR specifically were obtained from both the linear and the constrained libraries, proved by enzyme-linked immumosorbent assays and immunoprecipitation assays. DNA sequencing indicated that peptides displayed on some of the selected clones shared consensus sequences. Compared with traditional methods, this approach provided a cheaper and more rapid alternative to screen specific ligands for microcystin-LR. Moreover, since it is rather difficult to take small molecules as targets of phage display libraries, the success of this experiment expanded the applications of phage display technology, and provided a new avenue to study environmental small molecular toxins.

Expression modulation of multiple cytokines in vivo by cyanobacteria blooms extract from Taihu Lake, China.

Cyanobacterial blooms that generate microcystins (MCs) are being increasingly recognized as a potent health hazard in aquatic ecosystems. However, immunomodulation induced by cyanotoxins has not been well documented. This paper reports the in vivo data on the immune disorder caused by crude microcystin (MC) extract of cyanobacteria blooms collected from Taihu Lake, China, with respect to cytokine mRNA levels. Using reverse-transcriptional polymerase chain reaction (RT-PCR), the expression of multiple cytokines, including proinflammatory (IL-1beta, TNF-alpha, and IL-6) and Th1/Th2-related cytokines (IL-2, IL-4 and IL-10), was evaluated following the cyanobacteria blooms extract containing MCs (CBE) exposure at four doses of 23, 38, 77, 115 mg lyophilized algae cells/kg body weight. The results showed that the mRNA levels of TNF-alpha, IL-1beta, IL-2 and IL-4 decreased significantly following injection of all doses as compared to the control (LPS or ConA only), while the IL-6 level was unaffected. Contrast to this decrease, the level of IL-10 mRNA was, however, transiently up regulated following injection of the lowest dose of CBE. The distinct patterns of expression of these cytokines suggested a modulation of cytokine network, the essential component of the host immune system. We further developed a mathematical model to simulate the interaction of T helper cell subsets and related cytokines, which proved to be a good approach to study the kinetics of the interaction of cells and cytokines in microcystin immunosuppression.

Targeted gene silencing by small interfering RNA-based knock-down technology.

RNA interference (RNAi) has emerged as a powerful tool for the silencing of gene expression in animals and plants. RNAi is mediated by approximately 21-nt small interfering RNAs (siRNAs), which are originally produced from larger double stranded RNAs (dsRNAs) in vivo through the action of Dicer. Recently, many groups have reported systems designed to express siRNAs in mammalian cells through transfection of either oligonucleotides or plasmids encoding siRNAs. Although the use of siRNAs to silence genes in vertebrate cells was only reported three years ago, the emerging literature indicates that most vertebrate genes can be studied with this technology. This review summarizes some approaches to generate siRNAs, the delivery and application of siRNAs to target cells and the utility of siRNAs as analytical and potential therapeutic tools.

Analysis of microcystins in cyanobacteria blooms and surface water samples from Meiliang Bay, Taihu Lake, China.

Taihu Lake is the third largest freshwater lake in China. In recent years, the water pollution of cyanobacteria blooms has become a severe problem in this area. Microcystins (MCs) are an important group of toxic compounds mainly produced by some cyanobacteria species and have both acute and chronic hepatotoxic effects on animals and humans. This paper presents the first data on the identification and detection of MCs in both natural occurring cyanobacteria blooms and surface water samples (0-0.5 m), collected from Meiliang Bay, Taihu Lake, China. A conventional method for extraction and isolation of MCs from cyanobacteria blooms was applied. High-performance liquid chromatography (HPLC) analysis showed that the main toxic component in the cyanobacteria materials was MC-LR. The monoclonal antibody (mAb) against MC-LR produced by hybridoma technique was employed for direct competitive ELISA to detect the concentrations of MCs in bloom and water samples collected in 2001. The results not only revealed the presence of MCs but also temporal variations of MCs levels of three sampling stations in Meiliang Bay in 1 year. It is obvious that the MC contents were relatively higher during warm months and related with the status of eutrophication. Our study indicates the threat associated with MCs in water body of Taihu Lake. To prevent the MCs potential hazard on public health in this area, some necessary measures of monitoring and control of growth of cyanobacteria are urgently needed.

Effects of cyanobacteria bloom extract on some parameters of immune function in mice.

Microcystis aeruginosa is a common cyanobacterium in water blooms that appear world widely in eutrophic freshwaters, and its toxic blooms have caused many death and illness cases. This paper presents the first data on the immunotoxicity of a microcystin (MC) extract of cyanobacteria bloom collected from Taihu Lake, China to BALB/c mice. The cyanobacteria bloom extract (CBE) containing MCs was administered by i.p. injection for 14 days at three sublethal doses of 16, 32, 64 mg lyophilized algae cells/kg body weight. Exposure to CBE decreased body weights dose-dependently. Meanwhile, liver body weight ratios were markedly increased. The significant differences were also observed in spleen and thymus body ratios upon the elevation of treatment dose comparing to control. CBE was also found to reduce the phagocytosis evaluated using phagocytic index of peritoneal phagocyte; this suppression was not evident in percentage phagocytosis. Treatment of CBE produced the inhibition of lipopolysaccharide-induced lymphoproliferation and the dose-dependent decrease of the numbers of antibody-forming cells in mice that were immunized by using T-dependent antigen sheep red blood cells. However, CBE did not affect concanavalin A-induced T cell proliferation. Our results demonstrate that exposure to CBE resulted in immunosuppression in mice.

Expression of anti-neuroexcitation peptide (ANEP) of scorpion Buthus martensii Karsch in Escherichia coli.

According to the cDNA sequence of anti-neuroexcitation peptide of scorpion Buthus martensii Karsch, the putative mature anti-neuroexcitation peptide (ANEP) encoding DNA fragment was obtained by a PCR method, then was cloned into expression plasmid pET28a, fused with His tag at its 3' end. When expressed in E. coli BL21 (DE3), the expression of recombinant ANEP was 15% of total cellular proteins, while most recombinant ANEP products existed in the form of insoluble inclusion bodies. Coexpression of molecular chaperones or protein disulfide isomerase could not improve its solubility. The recombinant ANEP in the cell lysate was purified to homogeneity by metal chelating affinity chromatography and Superdex 30 chromatography. In bioassay with convulsive mice model induced by thiosemicarbazide, recombinant ANEP could apparently delay the convulsion seizure of model animals by 18% and showed anti-neuroexcitatory activity.

Expression and purification of recombinant human annexin V in Escherichia coli.

Human annexin V cDNA was cloned into plasmid pET19b and fused to a ten consecutive histidine tag at N-terminal. When expressed in E. coli BL21(DE3) LysS, the recombinant His10-annexin V accumulated in soluble form in the cytoplasm. By two-step chromatography, i.e., metal chelate affinity chromatography and anion exchange chromatography, recombinant His10-annexin V was purified to homogeneity on silver-stained SDS-PAGE gel. Recombinant annexin V, 7.4 mg, was obtained from a 1 litre flask culture.

Expression of human cardiac-specific homeobox protein in Escherichia coli.

Human cardiac-specific homeobox protein cDNA (hCsx) was cloned into expression plasmid pET32a and fused with Escherichia coli thioredoxin (Trx). The Trx-Csx fusion protein was under the control of bacteriophage T7 promoter. When expressed in E. coli BL21(DE3), about half of the recombinant Trx-Csx products existed in the form of insoluble inclusion bodies. When coexpressed with human protein disulfide isomerase, more than 90% of Trx-Csx products accumulated in the soluble form in the cell lysate. The recombinant Csx fusion protein was purified by one-step metal-chelating affinity chromatography.

[The manufacture and clinical use of the auto-sialographic instrument by remote controlling]

OBJECTIVE:The purpose of this article is to realize the auto-control of the injection of radiographic drugs in terms of the principle in remote controlling.METHODS:30 cases of sialaden diseases was studied using the auto sialographic instrument by remote controlling,compared to 67 cases studied using sialography by hand-controlling.RESULTS:The auto-sialographic instrument by remote controlling realize the telecontrol of the injection of the sialographic drugs and auto-injection,simplify the working procedure,and prevent medical staffs from X-ray radiation at work.CONCLUSION:The auto sialographic instrument by remote controlling can not only bring much convenience to medical staffs,but also improve the conformation to clinical diagnosis.