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1.
Purpose. The acidic microclimate in poly(D, L-lactide-co-glycolide) 50/50 microspheres has been previously demonstrated by our group as the primary instability source of encapsulated bovine serum albumin (BSA). The objectives of this study were to stabilize the encapsulated model protein, BSA, and to achieve continuous protein release by using a blend of: slowly degrading poly(D, L-lactide) (PLA), to reduce the production of acidic species during BSA release; and pore-forming poly(ethylene glycol) (PEG), to increase diffusion of BSA and polymer degradation products out of the polymer. Methods. Microspheres were formulated from blends of PLA (Mw 145,000) and PEG (Mw 10,000 or 35,000) by using an anhydrous oil-in-oil emulsion and solvent extraction (O/O) method. The polymer blend composition and phase miscibility were examined by FT-IR and DSC, respectively. Microsphere surface morphology, water uptake, and BSA release kinetics were also investigated. The stability of BSA encapsulated in microspheres was examined by losses in protein solubility, SDS-PAGE, IEF, CD, and fluorescence spectroscopy. Results. PEG was successfully incorporated in PLA microspheres and shown to possess partial miscibility with PLA. A protein loading level of 5% (w/w) was attained in PLA/PEG microspheres with a mean diameter of approximately 100 m. When PEG content was less than 20% in the blend, incomplete release of BSA was observed with the formation of insoluble, and primarily non-covalent aggregates. When 20%-30% PEG was incorporated in the blend formulation, in vitro continuous protein release over 29 days was exhibited. Unreleased BSA in these formulations was water-soluble and structurally intact. Conclusions. Stabilization and controlled relaease of BSA from PLA/PEG microspheres was achieved due to low acid and high water content in the blend formulation.  相似文献   

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Viral components can trigger autoimmunity, but the involved mechanisms remain to be elucidated. Toll-like receptor 3 (TLR3) recognizes viral double-stranded RNA (dsRNA) and appears to play an important role in this context. Our previous studies showed that signaling of TLR2, TLR3, TLR4 and TLR9 is highly redundant in the adjuvant effect needed to induce experimental autoimmune uveitis (EAU), an animal model of human autoimmune eye disease. In this study, we analyzed the effects of systemic delivery of polyinosinic:polycytidylic acid (poly(I:C)), a mimic of viral dsRNA, in the induction of EAU. We found that TLR3 agonist poly(I:C) enhanced EAU scores, DTH responses and Ag-specific T cell proliferation. In addition, Ag-specific Interleukin 17 (IL-17) and interferon gamma (IFN-γ) production by draining lymph node cells was markedly increased in the poly(I:C)-treated group. Our results suggest that activation of innate immune system mediated by TLR3 signaling pathway is of importance in the pathogenesis of virus-induced autoimmune diseases.  相似文献   

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在ip丝裂霉素C(MMC,5.20 mg·kg~(-1))前24h内将聚肌胞(polv I:C)sc给小鼠,可明显抑制MMC所诱发的小鼠骨髓细胞染色体畸变频率,具有剂量效应关系,Poly I:C(0.50 mg·kg~(-1))sc后4和12h对染色体畸变的抑制作用最强,而polvI:C(0.50 mg·kg~(-1))sc后4 h所取小鼠血清亦具有抑制染色体畸变的作用,结果提示polvI:C和其诱导的小鼠血清均可预防MMC对遗传物质的损伤作用。  相似文献   

5.

Background:

Impaired γ-aminobutyric acid (GABA) signaling may contribute to the emergence of cognitive deficits and subcortical dopaminergic hyperactivity in patients with schizophrenia and related psychotic disorders. Against this background, it has been proposed that pharmacological interventions targeting GABAergic dysfunctions may prove useful in correcting such cognitive impairments and dopaminergic imbalances.

Methods:

Here, we explored possible beneficial effects of the benzodiazepine-positive allosteric modulator SH-053-2’F-S-CH3, with partial selectivity at the α2, α3, and α5 subunits of the GABAA receptor in an immune-mediated neurodevelopmental disruption model. The model is based on prenatal administration of the viral mimetic polyriboinosinic-polyribocytidilic acid [poly(I:C)] in mice, which is known to capture various GABAergic, dopamine-related, and cognitive abnormalities implicated in schizophrenia and related disorders.

Results:

Real-time polymerase chain reaction analyses confirmed the expected alterations in GABAA receptor α subunit gene expression in the medial prefrontal cortices and ventral hippocampi of adult poly(I:C) offspring relative to control offspring. Systemic administration of SH-053-2’F-S-CH3 failed to normalize the poly(I:C)-induced deficits in working memory and social interaction, but instead impaired performance in these cognitive and behavioral domains both in control and poly(I:C) offspring. In contrast, SH-053-2’F-S-CH3 was highly effective in mitigating the poly(I:C)-induced amphetamine hypersensitivity phenotype without causing side effects in control offspring.

Conclusions:

Our preclinical data suggest that benzodiazepine-like positive allosteric modulators with activity at the α2, α3, and α5 subunits of the GABAA receptor may be particularly useful in correcting pathological overactivity of the dopaminergic system, but they may be ineffective in targeting multiple pathological domains that involve the co-existence of psychotic, social, and cognitive dysfunctions.  相似文献   

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Risperidone (RIS)-loaded microspheres based on poly(alkylene adipate)s derived from dicarboxylic acids and different aliphatic diols were prepared by the oil in water emulsion and solvent evaporation method. Specifically, 3 polyesters, namely poly(ethylene adipate), poly(propylene adipate), and poly(butylene adipate), were prepared with the aid of a 2-stage melt-polycondensation method and characterized by gel permeation chromatography, proton nuclear magnetic resonance (1H NMR), differential scanning calorimetry, and X-ray diffraction analysis. Results showed that the molecular weight of the polyesters increased as the diol molecular weight increased, while all polymers were of semi-crystalline nature and the melting temperature was varying from 49.1°C to 51.8°C and 65.9°C for poly(propylene adipate), poly(ethylene adipate), and poly(butylene adipate), respectively. The particle size of the RIS-loaded microspheres varied from 10 to 100 μm depending on the polyester type and the drug loading, while X-ray diffraction analysis revealed amorphous active pharmaceutical ingredient in the cases of high drug-loaded microspheres. In vitro drug release studies along with scanning electron microscopy images of microspheres after the completion of dissolution process showed that in all cases RIS release was controlled by the glass transition temperature of polyesters and physical state of active pharmaceutical ingredients via diffusion.  相似文献   

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Human nasal epithelium is an important physical barrier and innate immune defense protecting against inhaled substances and pathogens. Toll-like receptor (TLR) signaling, which plays a key role in the innate immune response, has not been well characterized in human nasal epithelial cells (HNECs), including the epithelial tight junctional barrier. In the present study, mRNAs of TLR1-10 were detected in hTERT-transfected HNECs, which can be used as an indispensable and stable model of normal HNECs, similar to primary cultured HNECs. To investigate the changes of tight junction proteins and the signal transduction pathways via TLRs in HNECs in vitro, hTERT-transfected HNECs were treated with TLR2 ligand P3CSK4, TLR3 ligand poly(I:C), TLR4 ligand LPS, TLR7/8 ligand CL097, TLR8 ligand ssRNA40/LyoVec, and TLR9 ligand ODN2006. In hTERT-transfected HNECs, treatment with poly(I:C) significantly reduced expression of the tight junction protein JAM-A and induced secretion of proinflammatory cytokines IL-8 and TNF-α. Both the reduction of JAM-A expression and the induction of secretion of IL-8 and TNF-α after treatment with poly(I:C) were modulated by distinct signal transduction pathways via EGFR, PI3K, and p38 MAPK and finally regulated by a TLR3-mediated NF-κB pathway. The control of TLR3-mediated signaling pathways in HNECs may be important not only in infection by viral dsRNA but also in autoimmune diseases caused by endogenous dsRNA released from necrotic cells.  相似文献   

9.
Purpose. This study describes the preparation and characterization of a controlled release formulation of granulocyte-macrophage colony-stimulating factor (GM-CSF) encapsulated in poly(glycolide-co-D,L-lactide) (PLGA) and poly(D,L-lactide) (PLA) microspheres. Methods. GM-CSF was encapsulated in PLGA/PLA microspheres by a novel silicone oil based phase separation process. Several different blends of PLGA and low molecular weight PLA were used to prepare the microspheres. The microspheres and the encapsulated GM-CSF were extensively characterized both in vitroand in vivo. Results. Steady release of GM-CSF was achieved over a period of about one week without significant 'burst' of protein from the microspheres. Analysis of microsphere degradation kinetics by gel permeation chromatography (GPC) indicated that low molecular weight PLA enhanced the degradation of the PLGA and thereby affected release kinetics. GM-CSF released from the microspheres was found to be biologically active and physically intact by bioassay and chromato-graphic analysis. Analysis of serum from mice receiving huGM-CSF indicated that the GM-CSF was biologically active and that a concentration of greater than 10 ng/mL was maintained for a period lasting at least nine days. MuGM-CSF was not detected followingin vivo administration of muGM-CSF microspheres. The tissues of mice receiving muGM-CSF microspheres were characterized by infiltration of neutrophils, and macrophages which were in significant excess of those found in mice administered with placebo controls (i.e. microspheres without GM-CSF). Conclusions. This study demonstrates the influence of formulation parameters on the encapsulation of GM-CSF in PLGA/PLA microspheres and its controlled release in biologically active form. The intense local tissue reaction in mice to muGM-CSF microspheres demonstrates the importance of the mode of delivery on the pharmacologic activity of GM-CSF.  相似文献   

10.
Experimental conditions necessary for the full expression of interferon-inducing activity by a complex of polyguanylic acid and polycytidylic acid included: (i) a sufficiently high molecular size of each homopolymer; (ii) annealing conditions which insured complete denaturation of polyguanylic acid self-structure; and (iii) the specific biological assay employed to assay interferon-inducing potency. The complex of polyguanylic acid and polycytidylic acid possessed several properties that suggested it may be an atypical polynucleotide interferon inducer. For instance, it was inactive in primary rabbit kidney cell cultures, usually exquisitely sensitive to polynucleotide interferon inducers, unless it was incubated on the cell cultures for prolonged times or in the presence of DEAE-dextran. Polyguanylic acid · polycytidylic acid could induce interferon in rabbits and mice but gave a more protracted response than did poly(I) · poly(C). Finally, poly(G) · poly(C) was, without any modification, resistant to degradation by serum nucleases.  相似文献   

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