Synthesis, pharmacophore modeling and in vitro activity of 10,11-dihydrodibenzo[b,f]oxepine-4-carboxamide derivatives as novel and potent antagonists of the prostaglandin EP4 receptor

The construction of a EP(4) antagonists pharmacophore model and the discovery of a highly potent oxepinic series of EP(4) antagonists is discussed. Compound 1a exhibits an excellent selectivity profile toward EP(2) receptor subtype and low cytochrome P450 inhibition potential.     

Targeting IL-15 receptor-bearing cells with an antagonist mutant IL-15/Fc protein prevents disease development and progression in murine collagen-induced arthritis

It has been suggested that the inflammatory cytokine IL-15 plays an important role in the development of several autoimmune diseases, including rheumatoid arthritis. We have generated a unique lytic and antagonistic IL-15 mutant/Fcgamma2a fusion protein (CRB-15) that targets the IL-15R. In the present study we examined the effects of targeting the IL-15R on the prevention and treatment of collagen-induced arthritis (CIA) in mice and probed the possible mechanisms of action of this IL-15 mutant/Fcgamma2a protein. Upon immunization with type II collagen, DBA/1 mice develop severe articular inflammation and destruction. Treatment of DBA/1 mice with a brief course of CRB-15 at the time of type II collagen challenge markedly inhibited the incidence and severity of arthritis. Moreover, in animals with ongoing established arthritis, treatment with CRB-15 effectively blocked disease progression compared with that in control-treated animals. The therapeutic effect of CRB-15 on either disease development or disease progression is remarkably stable, because withdrawal of treatment did not lead to disease relapse. A detailed analysis revealed that treatment with CRB-15 decreased synovitis in the joints; reduced bone erosion and cartilage destruction; reduced in situ production of the proinflammatory cytokines TNF-alpha, IL-1beta, IL-6, and IL-17; and decreased the responder frequency of autoreactive T cells. Our study suggests that the effective targeting of IL-15R-triggered events with CRB-15 can be of therapeutic importance in the treatment of rheumatoid arthritis.     

Gene doctoring: a method for recombineering in laboratory and pathogenic Escherichia coli strains

Background  

Homologous recombination mediated by the λ-Red genes is a common method for making chromosomal modifications in Escherichia coli. Several protocols have been developed that differ in the mechanisms by which DNA, carrying regions homologous to the chromosome, are delivered into the cell. A common technique is to electroporate linear DNA fragments into cells. Alternatively, DNA fragments are generated in vivo by digestion of a donor plasmid with a nuclease that does not cleave the host genome. In both cases the λ-Red gene products recombine homologous regions carried on the linear DNA fragments with the chromosome. We have successfully used both techniques to generate chromosomal mutations in E. coli K-12 strains. However, we have had limited success with these λ-Red based recombination techniques in pathogenic E. coli strains, which has led us to develop an enhanced protocol for recombineering in such strains.     

Epitope mapping of human thyroglobulin. Heterogeneous recognition by thyroid pathologic sera.

Thyroglobulin is the major Ag of the thyroid gland involved in autoimmune pathologies. Epitope mapping was carried out with a rabbit polyclonal immune serum against fusion proteins expressed in prokaryotic cells. After screening of an initial human thyroglobulin cDNA library and subcloning of immunoreactive clones, seven epitopes were characterized and localized on the human thyroglobulin monomeric molecule. One was close to each extremity of the molecule, and five others were concentrated in the middle, covering a sixth of this 2748-amino-acid chain. The immunoreactivities of 18 autoimmune sera from different thyroid pathologies were tested against the seven previously characterized epitopes. Those from Hashimoto's thyroiditis were the most immunoreactive. Immune responses were heterogeneous for sera from different pathologies as well as for those from the same pathology. The central epitopes and the near-C-terminal epitope, however, were the epitopes most often recognized by the immune sera. These findings show that some autoepitopes overlap accurately with some heteroepitopes characterized by a polyclonal immune serum directed against the mature protein.     

Polymorphism of the MHC class II Eb gene determines the protection against collagen-induced arthritis

Collagen-induced arthritis (CIA) is an animal model of auto immune polyarthritis, sharing similarities with rheumatoid arthritis (RA). Paradoxally, susceptibility to mouse CIA is controlled by the H2A loci (DQ homologous) while RA is linked to HLA.DR genes (H2E homologous). We recently showed that the E^d molecule prevents CIA development in susceptible H2 ^q mice. We addressed the question of whether H2Eb polymorphism will influence CIA incidence as HLA.DRB1 polymorphism does in RA. In F₁ mice, only H2Eb^d and H2Eb^s molecules showed protection. Using recombinant B10.RDD (Eb ^d/b) mice, we found that CIA protection was mediated by the first domain of the E^d molecule. Using peptides covering the third hypervariable region of the E chain, we found a perfect correlation between presentation of E peptides by the H2A^q molecule and protection on CIA. Therefore, the mechanism by which H2Eb protects against CIA seems to rely on the affinity of E peptides for the H2A_q molecule.     

Changes in Muscularis Propria of Anterior Vaginal Wall in Women with Pelvic Organ Prolapse

The objective of this study was to evaluate the morphological and immunohistochemical alterations of tissue removed from the upper third of anterior vaginal wall in a sample group of the female population presenting homogenous risk factors associated with pelvic organ prolapse (POP). The case study consisted of 14 patients with POP and there were 10 patients in the control group. Patient selection was carried on the basis of specific criteria and all of the patients involved in the study presented one or more of the recognized POP risk factors. Samples were taken from POP patients during vaginal plastic surgery following colpohysterectomy, and from control patients during closure of the posterior fornix following hysterectomy. Samples were processed for histological and immunohistochemical analyses for Collagen I and Collagen III, α-Smooth Muscle Actin (α-SMA), Platelet-Derived-Growth-Factor (PDGF), matrix metalloproteinase 3 (MMP3), tissue inhibitors metalloproteinase 1 (TIMP1), Caspase3. Immunofluorescence analyses for Collagen I and III and PDGF were also carried out. In prolapsed specimens our results show a disorganization of smooth muscle cells that appeared to have been displaced by an increased collagen III deposition resulting in rearrangement of the muscularis propria architecture. These findings suggest that the increase in the expression of collagen fibers in muscularis could probably be due to a phenotypic switch resulting in the dedifferentiation of smooth muscle cells into myofibroblasts. These alterations could be responsible for the compromising of the dynamic functionality of the pelvic floor.Key words: Pelvic organ prolapse, immunohistochemistry, smooth muscle cells, collagen, PDGF     

清醒猫摄食过程中的胃电活动

本文报告慢性清醒猫正常摄食过程中胃电的变化,以及脑室注射心得安和动物麻醉对胃电的影响。结果显示,空腹饥饿状态下,猫胃电图上出现具有特征性的高振幅的“饥饿波”;摄食时胃电慢波抑制,可见较小快波;进食后半小时左右,胃电开始出现每分钟4—5次的振幅逐渐增大的正弦形慢波,多数慢波负载有快波。脑室注射心得安,可使饱猫胃电慢波抑制期出现饥饿波。动物在饥饿时麻醉,胃电慢波幅度显著降低,饥饿波完全不出现,苏醒后逐渐恢复。     

Wheat inoculation with Azospirillum brasilense Sp6 and some mutants altered in nitrogen fixation and indole-3-acetic acid production

Abstract Inoculation of wheat seedlings with Azospirillum brasilense Sp6 produced an increase in the number and length of the lateral roots as a plant response. Inoculation with a Nif⁻ mutant, A. brasilense SpF103, which is producer of indole-3-acetic acid (IAA), yielded a very similar plant response. However, inoculation with a Nif⁻ mutant, A. brasilense SpF57, which is a low producer of IAA, did not elitic any response from the plant. The data suggest that the root system response of wheat seedlings to bacterial inoculation is due mainly to production of auxin-type substances by the microorganism.     

Modeling brewers' yeast flocculation

Flocculation of yeast cells occurs during the fermentation of beer. Partway through the fermentation the cells become flocculent and start to form flocs. If the environmental conditions, such as medium composition and fluid velocities in the tank, are optimal, the flocs will grow in size large enough to settle. After settling of the main part of the yeast the green beer is left, containing only a small amount of yeast necessary for rest conversions during the next process step, the lagering. The physical process of flocculation is a dynamic equilibrium of floc formation and floc breakup resulting in a bimodal size distribution containing single cells and flocs. The floc size distribution and the single cell amount were measured under the different conditions that occur during full scale fermentation. Influences on flocculation such as floc strength, specific power input, and total number of yeast cells in suspension were studied. A flocculation model was developed, and the measured data used for validation. Yeast floc formation can be described with the collision theory assuming a constant collision efficiency. The breakup of flocs appears to occur mainly via two mechanisms, the splitting of flocs and the erosion of yeast cells from the floc surface. The splitting rate determines the average floc size and the erosion rate determines the number of single cells. Regarding the size of the flocs with respect to the scale of turbulence, only the viscous subrange needs to be considered. With the model, the floc size distribution and the number of single cells can be predicted at a certain point during the fermentation. For this, the bond strength between the cells, the fractal dimension of the yeast, the specific power input in the tank and the number of yeast cells that are in suspension in the tank have to be known. Copyright 1998 John Wiley & Sons, Inc.