Liposomes and biotherapeutics

Application of liposomes as delivery system for biotherapeutic peptides and proteins may offer important therapeutic advantages over existing delivery methods. Several approaches towards achieving improved delivery of biotherapeutics with liposomes are outlined. Although the literature on this topic is sporadic and frequently incomplete, enough of a research foundation exists to justify the conclusion that liposomes can play an important role in the formulation and delivery of biotherapeutics. However, it will be necessary to understand more fully the mechanisms of action before optimum liposomal dosage forms can be designed.     

Local control of AMPA receptor trafficking at the postsynaptic terminal by a small GTPase of the Rab family

The delivery of neurotransmitter receptors into the synaptic membrane is essential for synaptic function and plasticity. However, the molecular mechanisms of these specialized trafficking events and their integration with the intracellular membrane transport machinery are virtually unknown. Here, we have investigated the role of the Rab family of membrane sorting proteins in the late stages of receptor trafficking into the postsynaptic membrane. We have identified Rab8, a vesicular transport protein associated with trans-Golgi network membranes, as a critical component of the cellular machinery that delivers AMPA-type glutamatergic receptors (AMPARs) into synapses. Using electron microscopic techniques, we have found that Rab8 is localized in close proximity to the synaptic membrane, including the postsynaptic density. Electrophysiological studies indicated that Rab8 is necessary for the synaptic delivery of AMPARs during plasticity (long-term potentiation) and during constitutive receptor cycling. In addition, Rab8 is required for AMPAR delivery into the spine surface, but not for receptor transport from the dendritic shaft into the spine compartment or for delivery into the dendritic surface. Therefore, Rab8 specifically drives the local delivery of AMPARs into synapses. These results demonstrate a new role for the cellular secretory machinery in the control of synaptic function and plasticity directly at the postsynaptic membrane.     

Vaginal infection with Ureaplasma urealyticum accounts for preterm delivery via induction of inflammatory responses

U. urealyticum, a member of the family Mycoplasmataceae, is often detected in the vagina of pregnant women. In this study, the possible association of ureaplasmal infection with preterm delivery was examined, as was the capacity of ureaplasmal LP to stimulate monocytes in vitro to produce pro-inflammatory cytokines relevant to preterm delivery. A hundred cases of normal delivery and 45 cases of preterm delivery were randomly selected. A mAb against U. urealyticum urease, that selectively and positively stained it in vaginal secretions of infected women but not in those of uninfected women, was generated. The preterm delivery group showed a significantly higher incidence of vaginal infection with this bacteria than the normal delivery group. Since the LP of Mycoplasma has potent biological activity, ureaplasmal LP was extracted. THP-1 cells, and human monocytic cells, produced IL-8, a potent pro-inflammatory cytokine associated with preterm delivery, and showed apoptotic cell death in response to the LP in vitro. These results suggest that U. urealyticum infection might play a causative role in preterm delivery via LP-induced IL-8 production and apoptosis.     

NgCAM and VAMP2 reveal that direct delivery and dendritic degradation maintain axonal polarity

Neurons are polarized cells of extreme scale and compartmentalization. To fulfill their role in electrochemical signaling, axons must maintain a specific complement of membrane proteins. Despite being the subject of considerable attention, the trafficking pathway of axonal membrane proteins is not well understood. Two pathways, direct delivery and transcytosis, have been proposed. Previous studies reached contradictory conclusions about which of these mediates delivery of axonal membrane proteins to their destination, in part because they evaluated long-term distribution changes and not vesicle transport. We developed a novel strategy to selectively label vesicles in different trafficking pathways and determined the trafficking of two canonical axonal membrane proteins, neuron–glia cell adhesion molecule and vesicle-associated membrane protein-2. Results from detailed quantitative analyses of transporting vesicles differed substantially from previous studies and found that axonal membrane proteins overwhelmingly undergo direct delivery. Transcytosis plays only a minor role in axonal delivery of these proteins. In addition, we identified a novel pathway by which wayward axonal proteins that reach the dendritic plasma membrane are targeted to lysosomes. These results redefine how axonal proteins achieve their polarized distribution, a crucial requirement for elucidating the underlying molecular mechanisms.     

Lectin functionalized nanocarriers for gene delivery

Gene therapy has emerged as one of the most promising therapeutic methods to treat various diseases. However, inadequate gene transfection efficacy during gene therapy demands further development of more efficient gene delivery strategies. Targeting genetic material to specific sites of action endows numerous advantages over non-targeted delivery. An ample variety of non-viral gene delivery vectors have been developed in recent years owing to the safety issues raised by viral vectors. Non-viral gene delivery vectors containing specific targeting ligands on their surfaces have been reported to enhance the gene transfection efficiency via receptor-mediated endocytosis for gene delivery. Among various targeting moieties investigated, carbohydrates and lectins (carbohydrate-binding proteins) played an essential role in gene delivery via either direct or reverse lectin targeting strategies. Lectins have a specific carbohydrate binding domain that can bind specifically to the carbohydrates. This review sheds light on various gene delivery nanovectors conjugated with either lectins or carbohydrates for enhanced gene transfection.     

In silico Structure Modeling and Comparative Analysis of Characterization Properties of Protein Polymers Useful for Protein-Based Nano Particulate Drug Delivery Systems (NPDDS): A Bioinformatics Approach

With an increasing interest in nanoparticulate delivery systems, there is a greater need to identify biomaterials that are biocompatible and safe for human applications. Protein polymers from animal and plant sources are promising materials for designing nanocarriers. Composition of the protein plays an important role for specific drug delivery applications such as drug release, targeting, and stimuli responsive drug release. An important issue in protein polymers is characteristics such as size, charge, and hydrophobicity may play a significant role in phagocytic uptake and initiating a subsequent immune response. This remains to be investigated systematically by analyzing factors that influence nanoparticle characteristics of protein and reduce phagocytic uptake and does not initiate immune response too. Although protein polymers are biodegradable, it is essential to ensure that there must not be premature enzymatic breakdown of the protein nanoparticles in the systemic circulation. Surface modification of the protein nanoparticles can be used to address this issue to propose the necessary modification in the surface of the protein would be great contribution in the nano particulate drug delivery systems (NPPDS). Of the various proteins, gelatin and albumin have been widely studied for drug delivery applications. Plant proteins are yet to be investigated widely for drug delivery applications so there is need to find out the plant proteins capable to act as nanoparticles. The commercial success of albumin-based nanoparticles has created an interest in other proteins. An increased understanding of the physicochemical properties coupled with the developments in rDNA technology will open up new opportunities for protein-based nanoparticulate systems. In the present studies several proteins currently useful for drug delivery system were structurally modeled and has been analyzed to propose the essential characteristics of protein for protein-based NPDDS.     

DESIGN OF HIGHLY EFFICIENT AND SELECTIVE TRANSFER REACTION OF NITROSYL GROUP TO dC AND dMC RESULTING IN SPECIFIC DEAMINATION

Nitric oxide (NO) is an important endogenous regulatory molecule, and S-nitrosothiols are believed to play a significant role in NO storage, transport, and delivery. Based on the ability to generate NO in vivo, S-nitrosothiols can be used as therapeutic drugs. In this study, we have developed an innovative method for sequence- and base-specific delivery of NO to a specific site of DNA followed by specific deamination.     

Design of highly efficient and selective transfer reaction of nitrosyl group to dc and d(M)C resulting in specific deamination

Nitric oxide (NO) is an important endogenous regulatory molecule, and S-nitrosothiols are believed to play a significant role in NO storage, transport, and delivery. Based on the ability to generate NO in vivo, S-nitrosothiols can be used as therapeutic drugs. In this study, we have developed an innovative method for sequence- and base-specific delivery of NO to a specific site of DNA followed by specific deamination.     

新的药物传递系统:外泌体作为药物载体递送*

外泌体(exosomes)是细胞分泌的囊泡,在细胞与细胞之间通信中发挥重要作用。由于其固有的长距离通信能力和出色的生物相容性而具有很大的潜力作为药物递送载体,尤其适合递送蛋白质、核酸、基因治疗剂等治疗药物。许多研究表明外泌体可以有效地将许多不同种类的货物递送至靶细胞,因此,它们常被作为药物载体用于治疗。对外泌体作为药物递送系统中面临的外泌体分离,药物装载和靶向治疗应用的进展与挑战作一介绍,以期更好为外泌体药物递送系统开发提供新思路。     

Survival of mycobacteria in macrophages is mediated by coronin 1-dependent activation of calcineurin

Pathogenic mycobacteria survive within macrophages by avoiding lysosomal delivery, instead residing in mycobacterial phagosomes. Upon infection, the leukocyte-specific protein coronin 1 is actively recruited to mycobacterial phagosomes, where it blocks lysosomal delivery by an unknown mechanism. Analysis of macrophages from coronin 1-deficient mice showed that coronin 1 is dispensable for F-actin-dependent processes such as phagocytosis, motility, and membrane ruffling. However, upon mycobacterial infection, coronin 1 was required for activation of the Ca(2+)-dependent phosphatase calcineurin, thereby blocking lysosomal delivery of mycobacteria. In the absence of coronin 1, calcineurin activity did not occur, resulting in lysosomal delivery and killing of mycobacteria. Furthermore, blocking calcineurin activation with cyclosporin A or FK506 led to lysosomal delivery and intracellular mycobacterial killing. These results demonstrate a role for coronin 1 in activating Ca(2+) dependent signaling processes in macrophages and reveal a function for calcineurin in the regulation of phagosome-lysosome fusion upon mycobacterial infection.     

In vitro interaction of the photoactive anticancer porphyrin derivative photofrin II with low density lipoprotein, and its delivery to cultured human fibroblasts

Low density lipoprotein (LDL) doped with the anticancer mixture of hematoporphyrin derivatives Photofrin II (P2) competes with native LDL for binding to fibroblast receptors, despite a slight increase in the negative net charge related to the presence of acidic residues of porphyrins. P2 delivery to fibroblasts can be achieved by LDL, HDL3 or albumin doped with P2 (LDL-P2, HDL-P2 or A-P2, respectively). P2 delivery to cells assessed by fluorescence measurement, is much more efficient, at low protein concentrations (10-20 micrograms/ml) by LDL-P2 than by HDL-P2 or A-P2. Moreover, P2 delivery to cells by LDL-P2 as a function of protein concentration is a saturable process, whereas P2 delivery by HDL-P2 or A-P2 is a linear process. Finally, reduction of the LDL-receptor number by preincubation of fibroblasts in medium supplemented with lipoproteins results in a decrease of P2 delivery by LDL-P2. These results suggest a special role of the LDL-receptor pathway in P2 delivery to cells and could be of interest in cancer phototherapy by porphyrins.     

Temporal integrity of an airborne odor stimulus is greatly affected by physical aspects of the odor delivery system

There is currently a debate about the role played by temporal patterns in neural activity in olfactory coding. An accurate analysis of this question, however, is only possible if the temporal properties of a stimulus itself are well defined. So far, no technique with sufficient temporal resolution has been available to accomplish this. Using a photoionization detector (PID), we show that the configuration of the odor delivery apparatus and the airflow settings greatly influence the integrity of a stimulus profile within an odor delivery apparatus. In a situation where pulsatile odor stimuli are applied to a stationary preparation, we tested the effect of 1) axial and off-center location within the airstream, 2) airflow of the odor delivery, 3) exit tube length, 4) exit tube diameter, 5) orientation of the odor delivery device in relation to the exhaust flow, and 6) exhaust tube air speed. This has important implications for the study of time in olfaction; significant planning must be incorporated into the design of the experiment to provide a well-defined odor delivery system.     

Injectable Lipid Emulsions—Advancements,Opportunities and Challenges

Injectable lipid emulsions, for decades, have been clinically used as an energy source for hospitalized patients by providing essential fatty acids and vitamins. Recent interest in utilizing lipid emulsions for delivering lipid soluble therapeutic agents, intravenously, has been continuously growing due to the biocompatible nature of the lipid-based delivery systems. Advancements in the area of novel lipids (olive oil and fish oil) have opened a new area for future clinical application of lipid-based injectable delivery systems that may provide a better safety profile over traditionally used long- and medium-chain triglycerides to critically ill patients. Formulation components and process parameters play critical role in the success of lipid injectable emulsions as drug delivery vehicles and hence need to be well integrated in the formulation development strategies. Physico-chemical properties of active therapeutic agents significantly impact pharmacokinetics and tissue disposition following intravenous administration of drug-containing lipid emulsion and hence need special attention while selecting such delivery vehicles. In summary, this review provides a broad overview of recent advancements in the field of novel lipids, opportunities for intravenous drug delivery, and challenges associated with injectable lipid emulsions.     

Protein disulfide-isomerase mediates delivery of nitric oxide redox derivatives into platelets

S-nitrosothiol compounds are important mediators of NO signalling and can give rise to various redox derivatives of NO: nitrosonium cation (NO+), nitroxyl anion (NO-) and NO* radical. Several enzymes and transporters have been implicated in the intracellular delivery of NO from S-nitrosothiols. In the present study we have investigated the role of GPx (glutathione peroxidase), the L-AT (L-amino acid transporter) system and PDI (protein disulfide-isomerase) in the delivery of NO redox derivatives into human platelets. Washed human platelets were treated with inhibitors of GPx, L-AT and PDI prior to exposure to donors of NO redox derivatives (S-nitrosoglutathione, Angeli's salt and diethylamine NONOate). Rapid delivery of NO-related signalling into platelets was monitored by cGMP accumulation and DAF-FM (4-amino-5-methylamino-2'7'-difluorofluorescein) fluorescence. All NO redox donors produced both a cGMP response and DAF-FM fluorescence in target platelets. NO delivery was blocked by inhibition of PDI in a dose-dependent manner. In contrast, inhibition of GPx and L-AT had only a minimal effect on NO-related signalling.PDI activity is therefore required for the rapid delivery into platelets of NO-related signals from donors of all NO redox derivatives. GPx and the L-AT system appeared to be unimportant in rapid NO signalling by the compounds used in the present study. This does not, however, exclude a possible role during exposure of cells to other S-nitrosothiol compounds, such as S-nitrosocysteine. These results further highlight the importance of PDI in mediating the action of a wide range of NO-related signals.     

微环DNA研究进展

质粒载体在基因治疗中占据重要地位.传统质粒DNA在真核生物中可能会引起严重的炎症反应,未甲基化的CpG序列可能抑制基因的表达,最好的解决办法是在生产质粒载体过程中将细菌调控序列整体消除.微环DNA是一种新颖的小环超螺旋表达框,它是传统质粒在大肠杆菌体内通过位点特异性重组得到的.微环DNA缺乏抗性标记基因、复制原点等细菌...     

Modeling bioaffinity-based targeted delivery of antimicrobials to Escherichia coli biofilms using yeast microparticles. Part I: Model development and numerical simulation

Biofilms are potential reservoirs for pathogenic microbes leading to a significant challenge for food safety, ecosystems, and human health. Various micro-and nanoparticles have been experimentally evaluated to improve biofilm inactivation by targeted delivery of antimicrobials. However, the role of transport processes and reaction kinetics of these delivery systems are not well understood. In this study, a mechanistic modeling approach was developed to understand the targeted delivery of chlorine to an Escherichia coli biofilm using a novel bioaffinity-based yeast microparticle. Biofilm inactivation by this delivery system was numerically simulated as a combination of reaction kinetics and transport phenomena. Simulation results demonstrate that the targeted delivery system achieved 7 log reduction within 16.2 min, while the equivalent level of conventional free chlorine achieved only 3.6 log reduction for the same treatment time. These numerical results matched the experimental observations in our previous study. This study illustrates the potential of a mechanistic modeling approach to improve fundamental understanding and guide the design of targeted inactivation of biofilms using biobased particles.     

Lipid and carbohydrate based adjuvant/carriers in immunology.

This review discusses various issues regarding vaccines; what are they and how they work, safety aspects, the role of adjuvants and carriers in vaccination, synthetic peptides as immunogens, and new technologies for vaccine development and delivery including the identification of novel adjuvants for mucosal vaccine delivery. There has been a recent increase of interest in the use of lipids and carbohydrates as adjuvants, and so a particular emphasis is placed on adjuvants derived from lipids or carbohydrates, or from both.     

Emotion enhances learning via norepinephrine regulation of AMPA-receptor trafficking

Emotion enhances our ability to form vivid memories of even trivial events. Norepinephrine (NE), a neuromodulator released during emotional arousal, plays a central role in the emotional regulation of memory. However, the underlying molecular mechanism remains elusive. Toward this aim, we have examined the role of NE in contextual memory formation and in the synaptic delivery of GluR1-containing alpha-amino-3-hydroxy-5-methyl-4-isoxazoleproprionic acid (AMPA)-type glutamate receptors during long-term potentiation (LTP), a candidate synaptic mechanism for learning. We found that NE, as well as emotional stress, induces phosphorylation of GluR1 at sites critical for its synaptic delivery. Phosphorylation at these sites is necessary and sufficient to lower the threshold for GluR1 synaptic incorporation during LTP. In behavioral experiments, NE can lower the threshold for memory formation in wild-type mice but not in mice carrying mutations in the GluR1 phosphorylation sites. Our results indicate that NE-driven phosphorylation of GluR1 facilitates the synaptic delivery of GluR1-containing AMPARs, lowering the threshold for LTP, thereby providing a molecular mechanism for how emotion enhances learning and memory.     

Targeting of herbal bioactives through folate receptors: a novel concept to enhance intracellular drug delivery in cancer therapy

Targeted drug delivery through folate receptor (FR) has emerged as a most biocompatible, target oriented, and non-immunogenic cargoes for the delivery of anticancer drugs. FRs are highly overexpressed in many tumor cells (like ovarian, lung, breast, kidney, brain, endometrial, and colon cancer), and targeting them through conjugates bearing specific ligand with encapsulated nanodrug moiety is undoubtedly, a promising approach toward tumor targeting. Folate, being an endogenous ligand, can be exploited well to affect various cellular events occurring during the progress of tumor, in a more natural and definite way. Thus, the aim of the review lies in summarizing the advancements taken place in the drug delivery system of different therapeutics through FRs and to refine its role as an endogenous ligand, in targeting of synthetic as well as natural bioactives. The review also provides an update on the patents received on the folate-based drug delivery system.     

Biglycan: an emerging small leucine-rich proteoglycan (SLRP) marker and its clinicopathological significance

Molecular and Cellular Biochemistry - Extracellular matrix (ECM) plays an important role in the structural organization of tissue and delivery of external cues to the cell. Biglycan, a class I...