The effects of proteoglycan surface patterning on neuronal pathfinding

Abstract

Protein micropatterning techniques are increasingly applied in cell choice assays to investigate fundamental biological phenomena that contribute to the host response to implanted biomaterials, and to explore the effects of protein stability and biological activity on cell behavior for in vitro cell studies. In the area of neuronal regeneration the protein micropatterning and cell choice assays are used to improve our understanding of the mechanisms directing nervous system during development and regenerative failure in the central nervous system (CNS) wound healing environment. In these cell assays, protein micropatterns need to be characterized for protein stability, bioactivity, and spatial distribution and then correlated with observed mammalian cell behavior using appropriate model system for CNS development and repair. This review provides the background on protein micropatterning for cell choice assays and describes some novel patterns that were developed to interrogate neuronal adaptation to inhibitory signals encountered in CNS injuries. Rationale for studying the role of proteoglycans in CNS injuriesNeurons from the central nervous system (CNS) possess a limited capacity to regenerate beyond scar tissue formation barriers in injuries even in the presence of minimal trauma to tissue organization [1,2]. A major culprit in CNS neuronal regenerative failure are the inihibitory proteoglycans (PGs), which are upregulated at the site of CNS injuries. PGs are composed of a core protein with varying numbers of attached glycosaminoglycan (GAG) side chains [3,4]. Proteoglycans are first translated intracellularly into proteins in the endoplasmic reticulum, and then GAG chains are later attached in the Golgi apparatus before secretion into the extracellular space [5][6][7][8]. The study of PGs has particularly been pronounced in the field of neurobiology and development of the nervous system. Numerous studies have concluded that PGs act as barriers that direct the migration of neural crest cells and the outgrowth direction of pathfinding neurons during development [9][10][11][12][13][14][15][16][17]. A number of these studies have additionally shown that cell guidance by PGs is based on an inhibitory signal located within the chondroitin sulfate GAG chains [9,18,19], with the result that the artificial addition of chondroitin sulfate (CS) sugars to the developing nervous system disrupts normal pathfinding [20] as well as does the removal of chondroitin sulfate chains through enzymatic treatment [9,19]. Davies et al. found that chondroitin sulfate proteoglycans (CSPG) are a major constituent of the glial scar and that dorsal root ganglion (DRG) outgrowth termination coincided with an increase in CSPG expression density [21]. Other sources have also shown that CSPGs are upregulated after CNS injuries [22][23][24][25].Interestingly, native adult CNS neurons actually posses the intrinsic ability to regenerate when the wound healing environment is prepared by eliminating inhibitory factors [23,26,27]. In additi...

Sugar glues for broken neurons

“…Previously, protein micro-patterning techniques have been utilized to create patterns for directing neuronal outgrowth. (193, 194) By utilizing an array of differentially sulfated GAG structures, one can create unique signaling combinations that enhance regeneration, functionality and directed growth of neurons.…”

Myelination and node of Ranvier formation on sensory neurons in a defined in vitro system

“…Myelination of neurons by Schwann cells has been extensively studied using dorsal root ganglia (DRG) cultures in a variety of serum containing and serum-free in vitro systems, with most systems relying on the use of biological substrates for cell culture (Callizot et al, 2011; Hlady and Hodgkinson, 2007; Liazoghli et al, 2011; Svenningsen et al, 2003; Wood et al, 1990). Less common has been node of Ranvier formation by Schwann cells on motoneurons (Pang et al, 2012; Rumsey et al, 2009) and by oligodendrocytes on CNS neurons (Pang et al, 2012) in in vitro systems.…”

Multiprotein Microcontact Printing with Micrometer Resolution

“…One such approach is to deposit precise spatial patterns of proteins on a material that serve to accomplish a biological goal. Recently, protein patterned surfaces have been used to control cell adhesion − and to direct the growth of neurons. − With the understanding that the cellular interactions with ECM are complex, a method that allows for the precise transfer of multiple independent protein patterns on the same surface becomes necessary. Despite advances toward this goal, no current techniques exist with the resolution necessary to pattern surfaces with multiple proteins at the cellular scale over a sufficiently large area.…”