Hemoglobin is an oxygen binding protein in erythrocytes. Hemoglobin is composed of four polypeptide chains. During the fetal stage the type of hemoglobin called fetal hemoglobin (HbF) dominates. After birth HbF is replaced by adult hemoglobin (HbA). HbF persists in concentrations less than 1%. Elevated concentration of HbF in adults exists in different conditions, Talassemi for example. When the uterus is damaged and the fetus doesn’t feel well its blood can pass the placenta barrier and enter the blood stream of the mother. A venous blood sample from the mother is analyzed to determine the status of the fetus. Laboratory Medicine Västernorrland already has two methods for analyzing HbF, one routine and one on call. The routine method needed to be replaced and the possibility to use flow cytometry was investigated. In this study, results from flow cytometry using Fetal Cell Count™ kit was compared to the results from the presently used methods, Kleihauer-Betke and HPLC. Cord blood was diluted with venous blood from an adult with the same blood group in various concentrations. A number of tests were performed and showed a fairly good correlation between the different methods. However more tests will be necessary to draw any clear conclusion.

Every year, several thousand individuals suffer spinal cord injury (SCI) in North America, while 1.5 million suffer traumatic brain injury in the U.S.A. alone. Primary mechanical trauma to the CNS is followed by a complex pathology, including vascular dysregulation, ischemia, edema and traumatic hemorrhage. Secondary damage is to a large extent caused by oxidative stress and inflammatory processes, resulting in necrosis and apoptosis of neural cells. If secondary tissue injury could be limited by interference with any of the pathomechanisms involved, preservation of structure and function would increase the potential for functional recovery. Experiments performed in other laboratories have shown that the polyphenolic flavonoid quercetin acts as an anti-oxidant and anti-inflammatory, reduces edema formation and apoptotic cell death. Quercetin is also an excellent iron chelator. This action profile suggested a high therapeutic potential for acute CNS trauma. Therefore, I used models of both spinal cord injury and head trauma in adult male rats to test the hypothesis that administration of quercetin is beneficial for the therapy of acute traumatic CNS injury. While the primary focus of my work was on therapy of acute traumatic spinal cord injury, quercetin was also evaluated in the settings of chronic SCI and acute head trauma. I found that, in a rat model of mid-thoracic spinal cord compression injury, 1) administration of quercetin, starting 1 hr after injury and continued every 12 hr, improved recovery of motor function in the hind limbs in more than half of the injured animals to a degree that allowed previously paraplegic animals to step or walk. The minimum quercetin dose that was efficacious was 5 µmol/kg. The minimum treatment duration for optimal outcome was determined to be 3 days. In control animals, some spontaneous recovery of motor function did occur, but never to an extent that allowed animals to step or walk. Quercetin administration was associated with more efficient iron clearance from the site of injury, decreased inflammatory response as reflected in decrease of myeloperoxidase activity and decreased apoptosis of neural cells at the site of injury. 2) Quercetin administered in the same injury model as late as 2 weeks after injury, given in a higher dose than that used for treatment in the acute phase, still resulted in significant recovery of motor function in 40% of the injured animals, although at a lower level of performance, when compared to early onset of treatment. 3) Quercetin administered after moderate fluid percussion brain injury resulted in decreased oxidative stress, as reflected in higher tissue glutathione levels at the site of injury. In animals receiving quercetin, the amplitude of compound action potentials was significantly better maintained at 24 hr and 72 hr after injury than in saline-treated control animals. My experiments have shown that the flavonoid quercetin is neuroprotective in a rat model of brain trauma and in a rat model of spinal cord injury. My data show that administration of quercetin after CNS trauma promotes iron clearance, decreases oxidative stress and inflammation. Quercetin also decreases apoptotic cell death following neurotrauma. These results suggest that quercetin may be a valuable adjunct in the therapy of acute CNS trauma. There is a possibility that administration of quercetin may be beneficial even in certain settings of chronic CNS trauma. These conclusions are based solely on the results from animal experiments. However, the fact that few adverse reactions have been noted to date in either animal experiments or human trials targeting other diseases is encouraging for the progression to human clinical trials for patients with spinal cord injury.

Trauma-induced spinal cord injury (SCI) is the most prevalent form of spinal cord injury affecting over 80% of the 36,000 Canadians living with this condition. The pathophysiological profile of traumatic SCI consists of an initial stage of direct damage followed by a series of secondary events, including reduced blood flow and increased generation of free radicals that leads to excitotoxicity, oxidative stress, hemorrhagic necrosis, inflammation, and apoptosis. We examined the hypotheses that delayed administration of the flavonoid quercetin inhibits the propagation of secondary events and promotes functional recovery after traumatic SCI by inhibiting inflammatory processes and signaling pathways that promote apoptosis and thereby promoting axon survival. To determine whether delayed quercetin treatment promoted functional recovery following SCI, male Wistar rats were subjected to a spinal cord compression injury by application of a 50 g modified aneurysm clip at the mid thoracic cord level. A treatment regimen of 75 µmol quercetin per kg rat or saline only (controls) was administered for a period of 3 days, 1 week or 2 weeks beginning at 2 weeks post surgery. Delayed quercetin treatment improved locomotion in injured animals although with severe deficit. To determine whether improved functional outcome correlated with improved tissue preservation and reduced scarring, we performed histological examinations at the injury site. In saline treated animals, at 8 weeks post injury we found over 80% of tissue loss with the majority of the remaining cells undergoing apoptosis. However, with 2 weeks delayed quercetin treatment, at least 50% of the tissue was still present at 8 weeks post surgery with a significant reduction of apoptosis. Quercetin treated animals also showed a reduction of reactive gliosis. To determine which intracellular signaling pathways may mediate the protective effects of quercetin, we carried out Western blots and immunocytochemical analyses of a number of potential pro-apoptotic pathways. We found that quercetin reduced the levels of the phosphorylated (activated) forms of the MAPK p38, ERK 1/2 (p42/44) and SAPK/JNK seen after SCI. We conclude that delayed quercetin treatment likely rescues neurons that would otherwise have died between the third and sixth weeks following injury by inhibiting apoptosis of glia cells. Quercetin may be acting via selective inhibition of kinase pathways that have been shown to be involved in apoptosis and cell growth. These findings not only reveal the protective effects of quercetin in reducing secondary damage after chronic SCI but also shed some light on some of the mechanisms underlying its actions.

This thesis describes the research, design, and development of a mid-drive wheelchair for use by the elders living independently, in assisted living facilities, and in nursing homes created by a design team at Georgia Tech's Center for Assistive Technology and Environmental Access (CATEA). This wheelchair stands to significantly improve the mobility of elders through better drive wheel placement and design features that could stand to improve their quality of life.

Objective: Ischemic spinal cord injury is a serious complication of aortic surgery. The mechanism underlying ischemic preconditioning (IPC) protection against spinal cord ischemia/reperfusion (I/R) injury is unclear. We investigated the role of spinal cord autoregulation in tolerance to spinal cord I/R injury induced by IPC. Although the extracellular signal-regulated kinases 1 and 2 (ERK1/2) are generally regarded as related to cell survival and proliferation, increasing evidence suggests that the role of the ERK1/2 pathway in I/R injury is contributory to inflammation. We investigated the effect of blocking ERK1/2 pathway to inhibit inflammation reaction in tolerance to spinal cord I/R injury. Methods: In the part 1 study, Sprague-Dawley rats were randomly assigned to 4 groups. IPC (P) group animals received IPC by temporary thoracic aortic occlusion (AO) with a 2-F Fogarty arterial embolectomy catheter for 3 min. I/R injury (I/R) group animals were treated with blood withdrawal and temporary AO for 12 min, and shed blood reinfusion at the end of the procedures. (P+I/R) group animals received IPC, followed by 5 min reperfusion, and then I/R procedures for 12 min. Sham (S) group animals received anesthesia and underwent surgical preparation only. Neurological functions were evaluated, and lumbar segments were harvested for histopathological examination. To evaluate the role of autoregulation in IPC, spinal cord blood flow and tissue oxygenation were continuously monitored throughout the procedure duration. In the part 2 study, spinal cord ischemia rats was induced by occluding the thoracic descending aorta with a balloon catheter introduced through a femoral artery, accompanied by concomitant exsanguinations. Rats in the control group were given dimethyl sulfoxide (vehicle) before undergoing spinal cord ischemia/reperfusion injury. In the U0126-treated group, rats were pretreated with an inhibitor of ERK1/2, U0126, to inhibit ERK1/2 phosphorylation. The sham rats underwent aortic catheterization without occlusion. Parameters, including neurologic status, neuronal survival, inflammatory cell infiltration, and interleukin-1£] production in the spinal cords, were compared between groups. Results: The Tarlov scores in the (I/R) group were significantly lower than those in the (S), (P), and (P+I/R) groups on days 1, 3, 5, and 7. The numbers of surviving motor neurons in the (S), (P), and (P+I/R) groups were significantly higher than those in the (I/R) group. The (P) group exhibited higher spinal cord blood flow and tissue oxygenation after reperfusion than the (S) group. The (P+I/R) group exhibited higher spinal cord blood flow and tissue oxygenation within the first 60 min after reperfusion than the (I/R) groups. In the part 2 study, early ERK1/2 phosphorylation was observed after injury in the control group, followed by abundant microglial accumulation in the infarct area and increased interleukin-1£] expression. In the U0126 group, U0126 treatment completely blocked ERK1/2 phosphorylation. Microglial activation and spinal cord interleukin-1£] levels were significantly reduced. Neuronal survival and functional performance were improved. Conclusions: IPC ameliorates spinal cord I/R injury in rats, probably mediated by triggering spinal cord autoregulation and improving local spinal cord blood flow and tissue oxygenation. The ERK1/2 pathway may play a noxious role in spinal cord ischemia/reperfusion injury by participating in inflammatory reactions and cytokine production. According to our findings, these concepts may be the new therapeutic targets in patients requiring aortic surgery.

A prototype of a one-time implantable spinal cord stimulation (SCS) system is presented in this thesis. A pair of inductive coils is used to achieve wireless power transmission and bidirectional communication. A rechargeable Li-ion battery is used to extend the lifetime of the implanted SCS device. Therefore, the number of the battery replacement surgery could be reduced such that one-time implantation is feasible. Besides, the proposed system on chip (SOC) controller and many discretes are integrated on a printed circuit board (PCB). The size of the proposed SCS device is competitive compared to the currently commercial products. The proposed SOC controller adopts a dual supply voltage scheme to reduce power consumption. The proposed SCS system employs an amplitude-shift keying (ASK) technique to carry out the data modulation and power transmission. One of the critical factors to affect efficiency of ASK-based wireless power transmission is the oscillating frequency accuracy. A ROM-less direct digital frequency synthesizer (DDFS) is presented in this thesis to fulfill such a high accuracy demand. Since the supply voltages of the discretes are diversified on a system PCB, many level converters are needed to translate different signal output voltage levels. To resolve above problem, the chip, then, must be redesigned to meet the various voltage level requirement, or added level convertors among the SOC and the discretes. Obviously, it will cause a lot of cost. A wide-range I/O buffer, thus, is proposed to resolve the compatibility problem caused by different supply voltages of discretes.

Spinal cord injury, acute pain, and morphine tolerance are important issues in the clinical practice. A primary injury to the spinal cord causes both morphological and biochemical changes with initiation of the devastating secondary pathophysiological pathways that ultimately destroy CNS cells and cause degeneration of nerve fibers. Tissue injury is associated with sensitization of nociceptors and subsequent changes in the excitability of central neurons, known as central sensitization. Nociceptor sensitization and central sensitization are believed to underlie the development of primary and secondary hyperalgesia, respectively. The most efficacious drugs used to relieve pain are the opioid analgesics. Chronic administration leads to the development of tolerance. Tolerance is manifested as a decreased potency of the drug, so that progressively larger doses must be administered to achieve a given level of analgesia. The processes underlying opioid tolerance still need to be elucidated. Recently, it is found calpain-Cdk5 (cyclin-dependent kinase-5)-p35 pathway modulation implicated in neuroprotection, acute nociceptive response, and morphine analgesia. In this thesis, we evaluate calpain inhibitor-MDL28170 and Cdk5 inhibitor-roscovitine against rat spinal cord hemisection, formalin-induced acute nociceptive responses, and chronic morphine tolerance. We found calpain-Cdk5-p35 pathway inhibition could protect spinal cord hemisection and subsequent neurodegeneration, inhibit formalin-induced flinch response involving DARPP-32 (dopamine and c-AMP regulated phosphoprotein, MW=32 kDa) phosphorylation, and reverse right shifted morphine dose-response curve with upregulated ED50 (50% of effective dose) reduction. Taken together, calpain-Cdk5-p35 pathway inhibition is useful in the management of spinal cord injury, acute inflammatory pain, and attenuate morphine tolerance development with further clinical application.

Abstract The first part of this research was focused on the relationship between injury severity and cell death mechanisms after spinal cord ischemia-reperfusion injury. The major blood supply to the thoracolumbar spinal cord comes from the segmental arteries originating from the thoracoabdominal aorta. Paraplegia cause by spinal cord ischemia is a devastating complication of thoracoabdominal aortic surgery. Previous studies indicated that ischemia-reperfusion injury of the central nervous system causes two distinct types of cell death, necrosis and apoptosis. It was also implicated that the intensity of injury can somehow affect the cell death mechanisms. In the first series of our experiments, by occluding the descending thoracic aorta with or without simultaneously inducing hypovolemic hypotension in rats, we established a model of experimental spinal cord ischemia-reperfusion (SCIR) in which the injury severity can be controlled. Recordings of carotid blood pressure (CBP) and spinal cord blood flow (SCBF) showed that aortic occlusion induced dramatic CBP elevation but SCBF drop in both the normotensive (NT) and hypotensive (HT) groups. However, the HT group demonstrated significantly lower SCBF during aortic occlusion, and much slower elevation of SCBF after reperfusion, and extremely poor neurological performance. Spinal cord lesions were characterized by infarction associated with extensive necrotic cell death, but little apoptosis and caspase-3 activity. In contrast, in the NT group, SCIR resulted in minor tissue destruction associated with persistently abundant apoptosis, augmented caspase-3 activity, and favorable functional outcome. The relative sparing of motoneurons in the ventral horns from apoptosis might have accounted for the minor functional impairment in the NT group. The severity of ischemia-reperfusion (I/R) injury was found to have substantial impact on the histopathological changes and cell death mechanisms, which correlated with neurological performance. These findings implicate that injury severity and duration after injury are two critical factors to be considered in therapeutic intervention. Based on the knowledge that bPrevious studies have implicated both excitotoxicity and apoptosis are involved in the pathogenesis of SCIR injury, we proposedtested the possibility that the N-methyl-D-aspartate (NMDA) receptor antagonist (dizocilpine maleate: (MK801) and the protein synthesis inhibitor (cycloheximide) would produce a synergic effect in the treatment of SCIR injury. In the second series of experiments, I/R iSpinal cord ischemia-reperfusion injury was induced by a thoracic aortic occlusion and blood volume reduction, followed by reperfusion and volume restoration. ischemia-reperfusion Rats were treated with vehicle, MK801, cycloheximide, or combination of MK801 and cycloheximide in combination. The MK801 and combined therapy group got a better recovery of hHind limb motor function recovery was better in the MK801 and combined-therapy groups than in the control and cycloheximide groups. On the 7th day after ischemia-reperfusion injury, all three treated groups showed significantly higher neuronal survival rates (NSR) than that of the control group. Among the three treated groups, the combined-treatment group showed the highest NSR. In addition, the Ttherapeutic effect of the combined-treatment group (27.4% increase of NSR) iwas better than the anticipated by the addition of MK801 and cycloheximide based on NSR data group. The number of apoptotic cells of was significantly reduced in the cycloheximide group and the combined-treatment group, as compared to that of the control group. It was unchanged in the MK-801 group. These results suggest that combined treatments directed at blocking both NMDA receptor-mediated excitotoxic necrosis and caspase-mediated apoptosis might have synergic therapeutic potential in reducing SCIR injury. Mitogen-activated protein kinases (MAPKs) including c-Jun N-terminal kinases (JNK), p38, and extracellular signal-regulated kinases (ERK), play important roles in the transduction of stressful signals and the integration of cellular responses. Although it has been generally held that the JNK and p38 pathways are related to cell death and degeneration, while the ERK pathway, cell proliferation and survival, controversy still exists. The roles of the ERK pathway in I/R injury of the CNS, in particular, remain to be clarified, because contradictory data have been reported by different investigators. Given this controversy, in the third series of experiments, we examined in injured spinal cords the temporal and spatial profiles of ERK1/2 activation following SCIR, and the effects of inhibiting the kinase that phosphorylates ERK1/2, MEK. The results showed that I/R injury induced an immediate phosphorylation of ERK1/2 in the spinal cord, which was alleviated by a MEK inhibitor, U0126. The control group was characterized by poorer neurological outcome, more severe tissue destruction, pronounced apoptosis, and lower neuronal survival. In contrast, the U0126-treated group demonstrated more apparent improvement of hind limb motor function, less tissue destruction, lack of apoptosis, and higher neuronal survival. In addition, administration of U0126 also significantly increased the activation of nuclear factor-£eB (NF-£eB) and the expression of cellular inhibitor of apoptosis protein 2 (c-IAP2). These findings implicate that the mechanisms underlying the neuroprotection afforded by ERK1/2 inhibition may be through the NF-£eB-c-IAP2 axis. The activation of the MEK-ERK signaling pathway appeared to be harmful in SCIR injury. Strategies aimed at blocking this pathway may bear potential therapeutic benefits in the treatment of SCIR injury. The second part of the research was focused on the pathophysiology of surgery-associated paraspinal muscle injury and measures to protect surgically violated paraspinal muscles. The wide dissection and forceful retraction of paraspinal muscles which are often required for posterior spinal sugery may severely jeopardize the muscles structurally and functionally. Immediate posteoperative pathological changes in the surgically violated paraspinal muscles may cause severe pain and a delay of patient ambulation. Long-term sequelae of surgical injury of paraspinal muscles include chronic back pain and impaired back muscle strength. Ironically, being a common complication of posterior spinal surgery, paraspinal muscle injury is so often neglected. Limited previous data indicate that the underlying pathophysiology of muscle damage involve both mechanical and ischemic mechanisms. We hypothesized that surgical dissection and retraction may produce oxidative stress within the paraspinal muscles. Meanwhile, we also proposed that the oxidative stress may trigger certain protective mechanisms within the insulted muscles. The first part of our study was a human study conducted to assess the significance of oxidative stress, and the relationship between it and the stress response mediated by heat shock protein 70 (HSP70) induction within paraspinal muscles under intraoperative retraction. A group of patients with lumbar spondylolisthesis treated with posterolateral lumbar spinal fusion, pedicle fixation and laminectomy were enrolled. Multifidus muscle specimens were harvested intraoperatively before, at designated time points during, and after surgical retraction. Muscle samples were analyzed for HSP70 and malondialdehyde (MDA) levels. Both HSP70 expression and MDA production within multifidus muscle cells were increased significantly by retraction. HSP70 expression then dropped after a peak at 1.5 hr of retraction, whereas MDA levels remained elevated even after release of retractors for reperfusion of the muscles. Histopathological and immunohistochemical evidence indicated that the decline of HSP70 synthesis within muscle cells after prolonged retraction was the result of severe muscle damage. These results highlighted the noxious impact of intraoperative retraction on human paraspinal muscles, and the significance of oxidative stress at the cellular and molecular levels. It is also implicated that intraoperative maneuvers aimed at reducing the oxidative stress within the paraspinal muscles may help attenuating surgery-associated paraspinal muscle damage. Given the findings of the first part of our study, and the knowledge that inflammation is a major postoperative pathological finding in surgically injured paraspinal muscles, we proceeded to examine the roles of two important inflammatory mediators, cyclooxygenase (COX)-2 and nuclear factor (NF)-£eB, in the pathogenesis of retraction-associated paraspinal muscle injury. A rat model of paraspinal muscle dissection and retraction that mimicks the conditions in human posterior spinal surgery was established. In the control group, paraspinal muscles were dissected from the spine through a dorsal incision, and then laterally retracted. Paraspinal muscle specimens were harvested before, and at designated time points during and after persistent retraction. The time course of NF-£eB activation as well as the expression of COX-2 were examined. Severity of inflammation was evaluated based on histopathology and myeloperoxidase (MPO) activity. NF-£eB activation was inhibited by the administration of pyrrolidine dithiolcarbamate (PDTC) in the PDTC-treated group. In the control group, retraction induced an early increase of NF-£eB/DNA binding activity in paraspinal muscle cells, which persited throughout the whole course of retraction. COX-2 expression was not detectable until 1 day after surgery, and reached a peak at 3 days. The time course of COX-2 expression correlated with that of inflammatory pathology and MPO activity. Extensive muscle fiber loss and collagen fiber replacement were observed at 7 days after surgery. Pretreatment with PDTC inhibited intraoperative NF-£eB activation and greatly downregulated postoperative COX-2 expression and inflammation in the muscles. Fibrosis following inflammation was also significantly abolished by PDTC administration. These findings indicate that NF-£eB-regulated COX-2 expression and inflammation play an important role in the pathogenesis of surgery-associated paraspinal muscle injury. Therapeutic strategies involving NF-£eB inhibition may be applicable to the prevention of such injury.

Thesis (Ph. D.)--Case Western Reserve University, 2004. / [School of Medicine] Department of Neurosciences. Includes bibliographical references. Available online via OhioLINK's ETD Center.

Diss. (sammanfattning) Stockholm : Karolinska institutet, 2009. / Härtill 4 uppsatser.