Toward modeling locomotion using electromyography-informed 3D models: application to cerebral palsy.

This position paper proposes a modeling pipeline to develop clinically relevant neuromusculoskeletal models to understand and treat complex neurological disorders. Although applicable to a variety of neurological conditions, we provide direct pipeline applicative examples in the context of cerebral palsy (CP). This paper highlights technologies in: (1) patient-specific segmental rigid body models developed from magnetic resonance imaging for use in inverse kinematics and inverse dynamics pipelines; (2) efficient population-based approaches to derive skeletal models and muscle origins/insertions that are useful for population statistics and consistent creation of continuum models; (3) continuum muscle descriptions to account for complex muscle architecture including spatially varying material properties with muscle wrapping; (4) muscle and tendon properties specific to CP; and (5) neural-based electromyography-informed methods for muscle force prediction. This represents a novel modeling pipeline that couples for the first time electromyography extracted features of disrupted neuromuscular behavior with advanced numerical methods for modeling CP-specific musculoskeletal morphology and function. The translation of such pipeline to the clinical level will provide a new class of biomarkers that objectively describe the neuromusculoskeletal determinants of pathological locomotion and complement current clinical assessment techniques, which often rely on subjective judgment. WIREs Syst Biol Med 2017, 9:e1368. doi: 10.1002/wsbm.1368 For further resources related to this article, please visit the WIREs website.

Activation of protein kinase C enhances peptide release from rat spinal cord slices.

One possible mechanism to account for the pain enhancing effects of protein kinase C (PKC) activation may be a facilitation of neurotransmitter release from terminals of nociceptive sensory neurons in the spinal cord. To examine this notion, we studied whether treatment with a phorbol ester enhanced the resting and capsaicin-evoked release of immunoreactive substance P (iSP) and immunoreactive calcitonin gene-related peptide (iCGRP) using an in vitro spinal cord slice preparation. Exposing the spinal cord tissue to 100 nM phorbol 12,13 dibutyrate (PDBu), an activator of PKC, results in a two-fold increase in the basal and the capsaicin-evoked release of iSP and iCGRF compared to evoked peptide release without PDBu. When the tissue was perfused with 1 microM 4-alpha PDBu, an analog of PDBu that does not activate PKC, the peptide release was not significantly different from control. Pre-exposing slices to 1 microM bisindolylmaleimide I, an inhibitor of PKC activity, prevents the facilitation of peptide release induced by PDBu. These results suggest that activation of PKC can augment the release of peptides in the spinal cord, which could increase nociceptive sensory transmission and contribute to hyperalgesia.

Activation of the epidermal platelet-activating factor receptor results in cytokine and cyclooxygenase-2 biosynthesis.

Recent studies suggest that the lipid mediator platelet-activating factor (PAF) is involved in keratinocyte function and skin inflammation. Indeed, PAF is found in association with inflammatory skin diseases, intradermal injections of PAF induce inflammation, and keratinocytes express functional PAF receptors (PAF-R). One mechanism by which the keratinocyte PAF-R could contribute to epidermal functions and inflammatory states would be through the synthesis of inflammatory regulators, such as PAF, PGs, and cytokines. The ability of the epidermal PAF-R to induce the synthesis of these immunomodulators was tested using a model system created by transduction of the PAF-R-negative human epidermal cell line KB with the PAF-R. Activation of this epidermal PAF-R resulted in arachidonic acid release, and the biosynthesis of PAF and PGE2. In addition, the KB PAF-R triggered increased levels of mRNA and protein for the inducible isozyme of cyclooxygenase (COX-2) as well as IL-6 and IL-8, both of which have been implicated in skin inflammatory processes. Studies with the human keratinocyte-derived epidermal cell line HaCaT revealed that activation of the endogenous PAF-R led to the increased accumulation of COX-2, IL-6, and IL-8 mRNA similar to that seen with the KB PAF-R model system. Finally, treatment of HaCaT keratinocytes with IL-8 resulted in PAF biosynthesis, indicating the existence of a positive feedback loop between IL-8 and PAF in epidermal cells. These studies suggest involvement of PAF and the PAF-R in the epidermal cytokine network.

Expression of the platelet-activating factor receptor results in enhanced ultraviolet B radiation-induced apoptosis in a human epidermal cell line.

Recent studies have demonstrated that ultraviolet B radiation (UVB) damages human keratinocytes in part by inducing oxidative stress and cytokine production. Severe UVB damage to the keratinocyte can also result in apoptosis or programmed cell death. Although the lipid mediator platelet-activating factor (PAF) is synthesized in response to epidermal cell damage and epidermal cells express PAF receptors, it is not known whether PAF is involved in UVB-induced epidermal cell apoptosis. These studies examined the role of the PAF system in UVB-induced epidermal cell apoptosis using a novel model system created by retroviral-mediated transduction of the PAF receptor-negative human epidermal cell line KB with the human PAF receptor (PAF-R). Expression of the PAF-R in KB cells did not affect base-line growth or apoptosis, yet resulted in a decrease in the lag time between treatment of the cells and the induction of apoptosis following irradiation with 400 J/m2 UVB. This effect was inhibited by pretreatment with the PAF-R antagonists WEB 2086 and A-85783, confirming involvement of the PAF-R in this process. At lower doses (100-200 J/m2) of UVB, only KB cells that expressed the PAF-R became apoptotic. Treatment of PAF-R-expressing KB clones with the metabolically stable PAF-R agonist 1-hexadexyl-2-N-methylcarbamoyl-3-glycerophosphocholine (CPAF) alone did not induce apoptosis but augmented the degree of apoptosis observed if CPAF was used in combination with lower doses (200 J/m2) of UVB irradiation. Interestingly, UVB irradiation was found to stimulate PAF synthesis only in PAF-R-expressing KB cell clones. The antioxidants N-acetyl cysteine, 1,1,3,3-tetramethyl-2-thiourea, and vitamin E inhibited both UVB-induced PAF biosynthesis as well as the augmentation of UVB-induced apoptosis in PAF-R-expressing KB clones, suggesting the possibility that UVB stimulates the production of oxidized lipid species with PAF-R agonistic activity in this model system. Thus, these studies indicate that a component of UVB-induced epidermal cell cytotoxicity can be modulated by PAF-R activation through the production of PAF and PAF-like species.

Identification and pharmacological characterization of platelet-activating factor and related 1-palmitoyl species in human inflammatory blistering diseases.

Through its pro-inflammatory effects on leukocytes, endothelial cells, and keratinocytes, the lipid mediator platelet-activating factor (PAF) has been implicated in cutaneous inflammation. Although the 1-alkyl PAF species has been considered historically the most abundant and important ligand for the PAF receptor (PAF-R), other putative ligands for this receptor have been described including 1-acyl analogs of sn-2 acetyl glycerophosphocholines. Previous bioassays have demonstrated a PAF-like activity in lesions of the autoimmune blistering disease bullous pemphigoid. To assess the actual sn-2 acetyl glycerophosphocholine species that result in this PAF agonistic activity, we measured PAF and related sn-2 acetyl GPCs in fresh blister fluid samples from bullous pemphigoid and noninflammatory (suction-induced) bullae by mass spectrometry. We report the presence of 1-hexadecyl as well as the 1-acyl PAF analog 1-palmitoyl-2-acetyl glycerophosphocholine (PAPC) in inflammatory blister fluid samples. Because PAPC is the most abundant sn-2 acetyl glycerophosphocholine species found in all samples examined, the pharmacological effects of this species with respect to the PAF-R were determined using a model system created by transduction of a PAF-R-negative epidermoid cell line with the PAF-R. Radioligand binding and intracellular calcium mobilization studies indicated that PAPC is approximately 100x less potent than PAF. Though a weak agonist, PAPC could induce PAF biosynthesis and PAF-R desensitization. Finally, intradermal injections of PAF and PAPC into the ventral ears of rats demonstrated that PAPC was 100x less potent in vivo. These studies suggest possible involvement of PAF and related species in inflammatory bullous diseases.

Activation of protein kinase C augments peptide release from rat sensory neurons.

To determine whether protein kinase C (PKC) mediates release of peptides from sensory neurons, we examined the effects of altering PKC activity on resting and evoked release of substance P (SP) and calcitonin gene-related peptide (CGRP). Exposing rat sensory neurons in culture to 10 or 50 nM phorbol 12,13-dibutyrate (PDBu) significantly increased SP and CGRP release at least 10-fold above resting levels, whereas the inactive 4alpha-PDBu analogue at 100 nM had no effect on release. Furthermore, 100 nM bradykinin increased peptide release approximately fivefold. Down-regulation of PKC significantly attenuated the release of peptides evoked by either PDBu or bradykinin. PDBu at 1 nM or 1-oleoyl-2-acetyl-sn-glycerol at 50 microM did not alter resting release of peptides, but augmented potassium- and capsaicin-stimulated release of both SP and CGRP approximately twofold. This sensitizing action of PKC activators on peptide release was significantly reduced by PKC down-regulation or by pretreating cultures with 10 nM staurosporine. These results establish that activation of PKC is important in the regulation of peptide release from sensory neurons. The PKC-induced enhancement of peptide release may be a mechanism underlying the neuronal sensitization that produces hyperalgesia.

Peptidase inhibitors improve recovery of substance P and calcitonin gene-related peptide release from rat spinal cord slices.

The purpose of present study was to determine whether peptidase activity affects the release of substance P (SP) and calcitonin gene-related peptide (CGRP) from spinal cord slices. When slices were exposed to various inhibitors of endopeptidase 24.11, the resting and capsaicin-stimulated release of SP were less than 0.04% and 0.20% total content per minute, respectively. Resting CGRP release was approximately 0.10% and stimulated release was 0.40%. The combination of 20 microM bacitracin, 100 microM phenylalanylalanine (Phe-Ala), and 50 microM p-chloromercuriphenylsulfonic acid (PCMS) significantly increased both resting and stimulated release of SP and CGRP at least two- or threefold. Doubling the concentration of PCMS and Phe-Ala did not further improve peptide release. These results demonstrate that recovery of peptides released from spinal cord slices is dependent in part on activity of multiple peptidases in the tissues.