Dictyostelium strains in which the gene encoding the cytoplasmic cAMP phosphodiesterase RegA is inactivated form small aggregates. This defect was corrected by introducing copies of the wild-type regA gene, indicating that the defect was solely the consequence of the loss of the phosphodiesterase. Using a computer-assisted motion analysis system, regA Ϫ mutant cells were found to show little sense of direction during aggregation. When labeled wild-type cells were followed in a field of aggregating regA Ϫ cells, they also failed to move in an orderly direction, indicating that signaling was impaired in mutant cell cultures. However, when labeled regA Ϫ cells were followed in a field of aggregating wild-type cells, they again failed to move in an orderly manner, primarily in the deduced fronts of waves, indicating that the chemotactic response was also impaired. Since wild-type cells must assess both the increasing spatial gradient and the increasing temporal gradient of cAMP in the front of a natural wave, the behavior of regA Ϫ cells was motion analyzed first in simulated temporal waves in the absence of spatial gradients and then was analyzed in spatial gradients in the absence of temporal waves. Our results demonstrate that RegA is involved neither in assessing the direction of a spatial gradient of cAMP nor in distinguishing between increasing and decreasing temporal gradients of cAMP. However, RegA is essential for specifically suppressing lateral pseudopod formation during the response to an increasing temporal gradient of cAMP, a necessary component of natural chemotaxis. We discuss the possibility that RegA functions in a network that regulates myosin phosphorylation by controlling internal cAMP levels, and, in support of that hypothesis, we demonstrate that myosin II does not localize in a normal manner to the cortex of regA Ϫ cells in an increasing temporal gradient of cAMP.

An ability to promote therapeutic immune cells to recognize cancer cells is important for the success of cell‐based cancer immunotherapy. We present a synthetic method for functionalizing the surface of natural killer (NK) cells with a supramolecular aptamer‐based polyvalent antibody mimic (PAM). The PAM is synthesized on the cell surface through nucleic acid assembly and hybridization. The data show that PAM has superiority over its monovalent counterpart in powering NKs to bind to cancer cells, and that PAM‐engineered NK cells exhibit the capability of killing cancer cells more effectively. Notably, aptamers can, in principle, be discovered against any cell receptors; moreover, the aptamers can be replaced by any other ligands when developing a PAM. Thus, this work has successfully demonstrated a technology platform for promoting interactions between immune and cancer cells.

No abstract

More than 90% of spinal cord injuries are caused by traumatic accidents and are often associated with other tissue damage (polytrauma) that can provide a source of continued pain input during recovery. In a clinically relevant spinal cord contusion injury model, prior work has shown that noxious stimulation at an intensity that engages pain (C) fibers soon after injury augments secondary injury and impairs functional recovery. Noxious input increases the expression of pro-inflammatory cytokines (interleukin 1β and 18), cellular signals associated with cell death (caspase 3 and 8), and physiological signs of hemorrhage. Here, it is shown that reducing neural excitability after spinal cord injury (SCI) with the local anesthetic lidocaine (micro-injected by means of a lumbar puncture) blocks these adverse cellular effects. In contrast, treatment with an analgesic dose of morphine had no effect. Contused rats that received nociceptive stimulation soon after injury exhibited poor locomotor recovery, less weight gain, and greater tissue loss at the site of injury. Prophylactic application of lidocaine blocked the adverse effect of nociceptive stimulation on behavioral recovery and reduced tissue loss from secondary injury. The results suggest that quieting neural excitability using lidocaine can reduce the adverse effect of pain input (from polytrauma or surgery) after SCI.

Pain (nociceptive) input caudal to a spinal contusion injury increases tissue loss and impairs long-term recovery. It was hypothesized that noxious stimulation has this effect because it engages unmyelinated pain (C) fibers that produce a state of over-excitation in central pathways. The present article explored this issue by assessing the effect of capsaicin, which activates C-fibers that express the transient receptor potential vanilloid receptor-1 (TRPV1). Rats received a lower thoracic (T11) contusion injury and capsaicin was applied to one hind paw the next day. For comparison, other animals received noxious electrical stimulation at an intensity that engages C fibers. Both forms of stimulation elicited similar levels of c-fos mRNA expression, a cellular marker of nociceptive activation, and impaired long-term behavioral recovery. Cellular assays were then performed to compare the acute effect of shock and capsaicin treatment. Both forms of noxious stimulation increased expression of tumor necrosis factor (TNF) and caspase-3, which promotes apoptotic cell death. Shock, but not capsaicin, enhanced expression of signals related to pyroptotic cell death [caspase-1, inteleukin-1 beta (IL-1ß)]. Pyroptosis has been linked to the activation of the P2X7 receptor and the outward flow of adenosine triphosphate (ATP) through the pannexin-1 channel. Blocking the P2X7 receptor with Brilliant Blue G (BBG) reduced the expression of signals related to pyroptotic cell death in contused rats that had received shock. Blocking the pannexin-1 channel with probenecid paradoxically had the opposite effect. BBG enhanced long-term recovery and lowered reactivity to mechanical stimulation applied to the girdle region (an index of chronic pain), but did not block the adverse effect of nociceptive stimulation. The results suggest that C-fiber input after injury impairs long-term recovery and that this effect may arise because it induces apoptotic cell death.

No abstract

Pain (nociceptive) input caudal to a spinal contusion injury can undermine long-term recovery and increase tissue loss (secondary injury). Prior work suggests that nociceptive stimulation has this effect because it fosters the breakdown of the blood-spinal cord barrier (BSCB) at the site of injury, allowing blood to infiltrate the tissue. The present study examined whether these effects impact tissue rostral and caudal to the site of injury. In addition, the study evaluated whether cutting communication with the brain, by means of a rostral transection, affects the development of hemorrhage. Eighteen hours after rats received a lower thoracic (T11–12) contusion injury, half underwent a spinal transection at T2. Noxious electrical stimulation (shock) was applied 6 h later. Cellular assays showed that, in non-transected rats, nociceptive stimulation increased hemoglobin content, activated pro-inflammatory cytokines and engaged signals related to cell death at the site of injury. These effects were not observed in transected animals. In the next experiment, the spinal transection was performed at the time of contusion injury. Nociceptive stimulation was applied 24 h later and tissue was sectioned for microscopy. In non-transected rats, nociceptive stimulation increased the area of hemorrhage and this effect was blocked by spinal transection. These findings imply that the adverse effect of noxious stimulation depends upon spared ascending fibers and the activation of rostral (brain) systems. If true, stimulation should induce less hemorrhage after a severe contusion injury that blocks transmission to the brain. To test this, rats were given a mild, moderate, or severe, injury and electrical stimulation was applied 24 h later. Histological analyses of longitudinal sections showed that nociceptive stimulation triggered less hemorrhage after a severe contusion injury. The results suggest that brain-dependent processes drive pain-induced hemorrhage after spinal cord injury (SCI).

No abstract