A relevant in vitro rat model for the evaluation of blood-brain barrier translocation of nanoparticles

Poly(MePEG₂₀₀₀cyanoacrylate-co-hexadecylcyanoacrylate) (PEG-PHDCA) nanoparticles have demonstrated their capacity to reach the rat central nervous system after intravenous injection. For insight into the transport of colloidal systems across the blood-brain barrier (BBB), we developed a relevant in vitro rat BBB model consisting of a coculture of rat brain endothelial cells (RBECs) and rat astrocytes. The RBECs used in our model displayed and retained structural characteristics of brain endothelial cells, such as expression of P-glycoprotein, occludin and ZO-1, and immunofluorescence studies showed the specific localization of occludin and ZO1. The high values of transendothelial electrical resistance and low permeability coefficients of marker molecules demonstrated the functionality of this model. The comparative passage of polyhexadecylcyanoacrylate and PEG-PHDCA nanoparticles through this model was investigated, showing a higher passage of PEGylated nanoparticles, presumably by endocytosis. This result was confirmed by confocal microscopy. Thanks to a good in vitro/in vivo correlation, this rat BBB model will help in understanding the mechanisms of nanoparticle translocation and in designing new types of colloidal carriers as brain delivery systems.Received 4 March 2005; accepted 14 April 2005     

Notch and cancer: a double-edged sword

The highly conserved Notch signaling pathway plays pleiotropic roles during embryonic development and is important for the regulation of selfrenewing tissues. The physiological functions of this signaling cascade range from stem cell maintenance and influencing cell fate decisions of barely differentiated progenitor cells, to the induction of terminal differentiation processes, all of which have been found to be recapitulated in different forms of cancers. Although Notch signaling has mostly been associated with oncogenic and growth-promoting roles, depending on the tissue type it can also function as a tumor suppressor. Here we describe recent findings on Notch signaling in cancer and tumor angiogenesis, and highlight some of the therapeutic approaches that are currently being developed to interfere with tumor growth and progression. Received 2 April 2007; received after revision 29 June 2007; accepted 2 July 2007     

Interferon receptors and their role in interferon action

Summary Interferon (IFN)_proteins interact with cells through specific cell surface receptors, some of which have been purified and cloned. The alpha-IFNs and beta-IFN bind to a common receptor (type I), whereas gamma-IFN binds to a separate receptor (type II). Both types of high-affinity receptors have been demonstrated on a variety of different kinds of cells but in relatively low numbers (10²–10⁴/cell). The relationship between IFN binding to receptors and the ways in which IFNs may affect cellular physiology and gene expression is discussed.     

The Janus kinase family of protein tyrosine kinases and their role in signaling

In the early 1990s, the search for protein kinases led to the discovery of a novel family of non-receptor tyrosine kinases, the Janus kinases or JAKs. These proteins were unusual because they contained two kinase homology domains and no other known signaling modules. It soon became clear that these were not ‘just another’ type of kinase. Their ability to complement mutant cells insensitive to interferons and to be activated by a variety of cytokines demonstrated their central signaling function. Now, as we approach the end of the decade, it is evident from biochemical studies to knockout mice that JAKs play non-redundant functions in development, differentiation, and host defense mechanisms. Here, recent progress is reviewed, with particular emphasis on structure-function studies aimed at revealing how this family of tyrosine kinases is regulated.     

Regulation of nicotinic acetylcholine receptors by tyrosine kinases in the peripheral and central nervous system: same players, different roles

Nicotinic acetylcholine receptors (nAChRs) exist in many subtypes and are found in the peripheral and central nervous system where they mediate or modulate synaptic transmission. We review how tyrosine phosphorylation and kinases regulate muscle and neuronal nAChRs. Interestingly, although some of the same kinase players interact with the various receptor subtypes, the functional consequences are different. While concerted action of MuSK, Abl- and Src-family kinases (SFKs) regulates the synaptic distribution of nAChRs at the neuromuscular junction, SFKs activate heteromeric neuronal nAChRs in adrenal chromaffin cells, thereby enhancing catecholamine secretion. In contrast, the activity of homomeric neuronal nAChRs, as found in the hippocampus, is negatively regulated by tyrosine phosphorylation and SFKs. It appears that tyrosine kinases provide the means to regulate all nAChRs; but the functional consequences, even those caused by the same kinase family, are specific for each receptor subtype and location. Received 21 February 2006; received after revision 24 July 2006; accepted 30 August 2006     

Role of dystrophin and utrophin for assembly and function of the dystrophin glycoprotein complex in non-muscle tissue

The dystrophin glycoprotein complex (DGC) is a multimeric protein assembly associated with either the X-linked cytoskeletal protein dystrophin or its autosomal homologue utrophin. In striated muscle cells, the DGC links the extracellular matrix to the actin cytoskeleton and mediates three major functions: structural stability of the plasma membrane, ion homeostasis, and transmembrane signaling. Mutations affecting the DGC underlie major forms of congenital muscle dystrophies. The DGC is prominent also in the central and peripheral nervous system and in tissues with a secretory function or which form barriers between functional compartments, such as the blood-brain barrier, choroid plexus, or kidney. A considerable molecular heterogeneity arises from cell-specific expression of its constituent proteins, notably short C-terminal isoforms of dystrophin. Experimentally, the generation of mice carrying targeted gene deletions affecting the DGC has clarified the interdependence of DGC proteins for assembly of the complex and revealed its importance for brain development and regulation of the ’milieu intérieur. Here, we focus on recent studies of the DGC in brain, blood-brain barrier and choroid plexus, retina, and kidney and discuss the role of dystrophin isoforms and utrophin for assembly of the complex in these tissues. Received 4 October 2005; received after revision 14 March 2006; accepted 5 April 2006     

Molecular guardians for newborn proteins: ribosome-associated chaperones and their role in protein folding

A central dogma in biology is the conversion of genetic information into active proteins. The biosynthesis of proteins by ribosomes and the subsequent folding of newly made proteins represent the last crucial steps in this process. To guarantee the correct folding of newly made proteins, a complex chaperone network is required in all cells. In concert with ongoing protein biosynthesis, ribosome-associated factors can interact directly with emerging nascent polypeptides to protect them from degradation or aggregation, to promote folding into their native structure, or to otherwise contribute to their folding program. Eukaryotic cells possess two major ribosome-associated systems, an Hsp70/Hsp40-based chaperone system and the functionally enigmatic NAC complex, whereas prokaryotes employ the Trigger Factor chaperone. Recent structural insights into Trigger Factor reveal an intricate cradle-like structure that, together with the exit site of the ribosome, forms a protected environment for the folding of newly synthesized proteins. Received 29 June 2005; received after revision 4 August 2005; accepted 18 August 2005     

Prolactin inducible protein in cancer, fertility and immunoregulation: structure, function and its clinical implications

Prolactin inducible protein (PIP) is a 17- kDa single polypeptide chain, known by various names due to its versatile nature and function in human reproductive and immunological systems. It is expressed in several exocrine tissues such as the lacrimal, salivary, and sweat glands. Its expression is up regulated by prolactin and androgens, and estrogens down regulate it. Due to its over-expression in metastatic breast and prostate cancer, presently PIP is considered as a prognostic biomarker. Moreover, its aspartyl-proteinase nature suggests its role in tumor progression. PIP has unique features because it is small in size and plays multiple important functions. Its ability to bind potentially with CD4-T cell receptor, immunoglobulin G (IgG), actin, zinc α2-glycoprotein (ZAG), fibronectin and enamel pellicle, reveals its important biological functions. This is the first comprehensive review on the structure and functional analysis of PIP and its clinical applications. Received 04 August 2008; received after revision 09 September 2008; accepted 15 September 2008     

Myelin sheaths: glycoproteins involved in their formation,maintenance and degeneration

Myelin sheaths are formed around axons by extending, biochemically modifying and spiraling plasma membranes of Schwann cells in the peripheral nervous system (PNS) and oligodendrocytes in the central nervous system (CNS). Because glycoproteins are prominent components of plasma membranes, it is not surprising that they have important roles in the formation, maintenance and degeneration of myelin sheaths. The emphasis in this review is on four integral membrane glycoproteins. Two of them, protein zero (P0) and peripheral myelin protein-22 (PMP-22), are components of compact PNS myelin. The other two are preferentially localized in membranes of sheaths that are distinct from compact myelin. One is the myelin-associated glycoprotein, which is localized at the inside of sheaths where it functions in glia-axon interactions in both the PNS and CNS. The other is the myelin-oligodendrocyte glycoprotein, which is preferentially localized on the outside of CNS myelin sheaths and appears to be an important target antigen in autoimmune demyelinating diseases such as multiple sclerosis. Received 8 April 2002; received after revision 13 May 2002; accepted 22 May 2002     

Endogenous synthesis of peptidoglycan in eukaryotic cells; a novel concept involving its essential role in cell division,tumor formation and the biological clock

Degradation products of peptidoglycan, the universal bacterial cell wall constituent, were previously found in animal tissues and urine. Reassessment and quantitative analysis of available data lead to an original concept, i.e. that eukaryotic cells synthesize peptidoglycan. We present a model in which this endogenously synthesized peptidoglycan is essential for the processes of eukaryotic cell division and sleep induction in animals. Genes for peptidoglycan metabolism, like those for lysine biosynthesis in plants, are probably inherited from endosymbiotic bacteria, the ancestors of mitochondria and chloroplasts. Corollaries of this concept, i.e. roles for peptidoglycan metabolism in tumor formation and in the biological clock, are supported by abundant evidence. We propose that many interactions between bacteria and eukaryotes are conditioned by their common genetic heritage.     

Differential binding of conA and WGA on the cell surface,the role of sialic acid in their expression and the increased activity of sialidase after cis-Platin treatment

Summary It is reported that concanavalin A (conA) and wheat germ agglutinin (WGA) have a differential binding pattern on normal mouse spleen lymphocytes and the surface of Dalton's lymphoma cells. It is suggested that sialic acid on the cell surface controls the expression of lectin binding sites. Further, it has been observed that the increased release of sialic acid from cell surfaces aftercis-dichlorodiammine platinum (II) (cis-Platin) treatment is due to the increased activity of sialidase.To whom reprint requests should be addressed.     

Towards an understanding of the role of glutamate in neurodegenerative disorders: energy metabolism and neuropathology

It is thought that impairment, of energy metabolism that results in deterioration of membrane function, leading to loss of the Mg²⁺ block on NMDA receptors, and allowing persistent activation of these receptors by glutamate, might be a cause of neuronal death in neurodegenerative disorders. Studies in rodents using mitochondrial respiratory chain toxins, such as aminooxyacetic acid, 1-methyl-4-phenylpyridinium, malonic acid and 3-nitropropionic acid, suggest that such processes may indeed be involved in neurotoxicity. Striatal and nigral degeneration induced by mitochondrial toxins in rodents resembles the neuropathology seen in humans suffering from Huntington's or Parkinson's disease, and can be prevented either by decortication or by NMDA receptor antagonists. Such experimental observations suggest that glutamate may be involved in neuronal death leading to neurodegenerative disorders in humans. If so, glutamate antagonists may offer a therapeutic approach for retarding the progression of these disabling disorders.     

N-acetylmuramic acid 6-phosphate lyases (MurNAc etherases): role in cell wall metabolism, distribution, structure, and mechanism

MurNAc etherases cleave the uniqued-lactyl ether bond of the bacterial cell wall sugar N-acetylmuramic acid (MurNAc). Members of this newly discovered family of enzymes are widely distributed among bacteria and are required to utilize peptidoglycan fragments obtained either from the environment or from the endogenous cell wall (i.e., recycling). MurNAc etherases are strictly dependent on the substrate MurNAc possessing a free reducing end and a phosphoryl group at C6. They carry a single conserved sugar phosphate isomerase/sugar phosphate- binding (SIS) domain to which MurNAc 6-phosphate is bound. Two subunits form an enzymatically active homodimer that structurally resembles the isomerase module of the double-SIS domain protein GlmS, the glucosamine 6-phosphate synthase. Structural comparison provides insights into the two-step lyase-type reaction mechanism of MurNAc etherases: β-elimination of the D-lactic acid substituent proceeds through a 2,3-unsaturated sugar intermediate to which water is subsequently added. Received 31 August 2007; received after revision 12 October 2007; accepted 1 November 2007