MiRNAs and inflammatory bowel disease: An interesting new story

Inflammatory bowel disease (IBD), as a chronic and recurrent inflammatory disorder, is caused by a dysregulated and aberrant immune response to exposed environmental factors in genetically susceptible individuals. Despite huge efforts in determining the molecular pathogenesis of IBD, an increasing worldwide incidence of IBD has been reported. MicroRNAs (miRNAs) are a set of noncoding RNA molecules that are about 22 nucleotides long, and these molecules are involved in the regulation of the gene expression. By clarifying the important role of miRNAs in a number of diseases, their role was also considered in IBD; numerous studies have been performed on this topic. In this review, we attempt to summarize a number of studies and discuss some of the recent developments in the roles of miRNAs in the pathophysiology, diagnosis, and treatment of IBD.     

Halothiophene benzimidazoles as P1 surrogates of inhibitors of blood coagulation factor Xa

Neutral weak halothiophene benzimidazole inhibitors of the serine protease factor Xa were identified via screening of a compound library. The X-ray crystal structure of representative 3a bound to human fXa confirmed the S1 binding mode. Starting from 3a a series of halothiophene benzimidazoles was synthesized and investigated for their factor Xa inhibitory activity. This led to potent and selective achiral inhibitors against fXa such as compounds 9k and 9w.     

Modification of the Stewart biphasic colorimetric assay for stable and accurate quantitative determination of Pluronic and Tetronic block copolymers for application in biological systems

Block copolymers are increasingly being applied in areas such as transfection, membrane sealing, site-specific targeting, and bionanoengineering yet there is a relative paucity of assays available for simple, stable and reproducible colorimetric determination of copolymer concentration in solution. We have focused on improving the accuracy and reproducibility of a modified version of the Stewart biphasic colorimetric assay for quantitative determination of Pluronic (poloxamer) and Tetronic (poloxamine) macromolecules. The optimized assay achieved linear response ranges in chloroform for commonly used copolymers such as poloxamine 904 (20-300 microg/ml), poloxamine 908 (10-400 microg/ml), poloxamer 402 (20-400 microg/ml), and poloxamer 407 (10-400 microg/ml). Variation in the type of chlorinated solvent used significantly increased assay sensitivity, presumably through macromolecular reorientation, affording increased access for copolymer-ferrothiocyanate complexation. This was found to be optimally favored by the planar geometry of the solvent cis 1,2-dichloroethylene. For application to biological systems copolymer-protein interactions were for the first time determined and were found to be dependent on the fraction of hydrophobic constituents of the block copolymers and protein type. For instance serum albumin was found to interact with copolymers of low hydrophilic-lipophilic balance values and poly(propylene oxide) contaminants, whereas this interaction was not significant with the relatively hydrophilic IgG. In such systems the colorimetric assay directly determines the fraction of unbound (free) copolymer in the presence of proteins.     

Exposure to Leishmania major modulates the proportion of CD4+ T cells without affecting cellular immune responses

The immune responses of individuals exposed to Leishmania major were evaluated and compared with those of non-exposed volunteers. Forty-one patients with active lesion(s), 43 healed individuals, 15 vaccinees 1 month or 1 year post vaccination, and 15 non-exposed volunteers were studied. Leishmanin skin test (LST) response, proliferative response of lymphocyte (PRL) to L. major antigen, IFN-gamma and IL-4 production, and percentage of L. major-specific CD4+, CD8+ and CD16+/CD56+ cells in peripheral blood mononuclear cells were assessed. Data showed positive LST (>5 mm) in 92% of patients, 98% of healed, and 80% or 43% of vaccinees 1 month and 1 year post vaccination, respectively. Positive PRL (SI>2.5) was displayed in 90%, 84%, 46% and 7% of patients, healed, vaccinated (post 1 year) and non-exposed donors, respectively. The mean +/-S.E. of IFN-gamma was 924 +/- 149, 1,278 +/- 185, 470 +/- 282 or 258 +/- 82 pg/ml in patients, healed cases and vaccinees after 1 month or 1 year, respectively. Positive IFN-gamma responders (>300 pg/ml) were shown in 72% of patients, 81% of healed cases, 31% or 39% of vaccinees and 0% of non-exposed donors. A reduced percentage of CD4+ T-cells and an increased percentage of NK cells were found in exposed individuals compared to non-exposed donors. The data indicated that exposure to L. major modulates the proportion of CD4+ T cells and increases NK cells percentage. However, the cellular immune responses including induction of LST, and IFN-gamma production are increased in exposed individuals.     

Molecularly imprinted polymer cartridges coupled on-line with high performance liquid chromatography for simple and rapid analysis of dextromethorphan in human plasma samples

In this paper, a novel method is described for automated determination of dextromethorphan in biological fluids using molecularly imprinted solid-phase extraction (MISPE) as a sample clean-up technique combined with high performance liquid chromatography (HPLC). The water-compatible molecularly imprinted polymers (MIPs) were prepared using methacrylic acid as functional monomer, ethylene glycol dimethacrylate as cross-linker, chloroform as porogen and dextromethorphan as template molecule. These imprinted polymers were used as solid-phase extraction sorbent for the extraction of dextromethorphan from human plasma samples. Various parameters affecting the extraction efficiency of the MIP cartridges were evaluated. The high selectivity of the sorbent coupled to the high performance liquid chromatographic system permitted a simple and rapid analysis of this drug in plasma samples with limits of detection (LOD) and quantification (LOQ) of 0.12 ng/mL and 0.35 ng/mL, respectively. The MIP selectivity was evaluated by analyzing of the dextromethorphan in presence of several substances with similar molecular structures and properties. Results from the HPLC analyses showed that the recoveries of dextromethorphan using MIP cartridges from human plasma samples in the range of 1-50 ng/mL were higher than 87%.     

Reduction of spontaneous metastases through induction of carbohydrate cross-reactive apoptotic antibodies

The selective targeting of tumor-associated carbohydrate Ags by the induction of serum Abs that trigger apoptosis of tumor cells as a means to reduce circulating tumor cells and micrometastases would be an advantage in cancer vaccine development. Some plant lectins like Griffonia simplicifolia lectin I and wheat germ agglutinin mediate the apoptosis of tumor cells. We investigated the possibility of using these lectins as templates to select peptide mimotopes of tumor-associated carbohydrate Ags as immunogens to generate cross-reactive Abs capable of mediating apoptosis of tumor cells. In this study, we show that immunization with a mimotope selected based on its reactivity with Griffonia simplicifolia lectin I and wheat germ agglutinin induced serum IgM Abs in mice that mediated the apoptosis of murine 4T1 and human MCF7 cell lines in vitro, paralleling the apoptotic activity of the lectins. Vaccine-induced anti-carbohydrate Abs reduced the outgrowth of micrometastases in the 4T1 spontaneous tumor model, significantly increasing survival time of tumor-bearing animals. This finding parallels suggestions that carbohydrate-reactive IgM with apoptotic activity may have merit in the adjuvant setting if the right carbohydrate-associated targets are identified.     

DNA binding studies of 3, 5, 6-trichloro-2-pyridinol pesticide metabolite

3, 5, 6-Trichloro-2-pyridinol (TCP) is a stable metabolite of two major pesticides, Chlopyrifos insecticide and Triclopyr herbicide, which are widely used in the world. The potential health hazard associated with TCP is identified due to its high affinity to the DNA molecule. Therefore, in this study, the interaction of native calf thymus DNA with TCP has been investigated using spectrophotometric, circular dichroism (CD), spectrofluorometric, viscometric and voltametric techniques. It was found that TCP molecules could interact with DNA via a groove-binding mode, as evidenced by hyperchromism, with no red shift in the UV absorption band of TCP, no changes in K(b) values in the presence of salt, no significant changes in the specific viscosity and CD spectra of DNA, and a decrease in peak currents with no shift in the voltamogram. In addition, TCP is able to release Hoechst 33258, a strong groove binder, in the DNA solutions. The results are indicative of the groove-binding mode of TCP to DNA.