Primordial germ cells (PGCs) share many properties with embryonic stem cells (ESCs) and innately ... more Primordial germ cells (PGCs) share many properties with embryonic stem cells (ESCs) and innately express several key pluripotency-controlling factors, including OCT4, NANOG, and LIN28. Therefore, PGCs may provide a simple and efficient model for studying somatic cell reprogramming to induced pluripotent stem cells (iPSCs), especially in determining the regulatory mechanisms that fundamentally define pluripotency. Here, we report a novel model of PGC reprogramming to generate iPSCs via transfection with SOX2 and OCT4 using integrative lentiviral. We also show the feasibility of using non-integrative approaches for generating iPSC from PGCs using only these two factors. We show that human PGCs express endogenous levels of KLF4 and C-MYC protein at levels similar to embryonic germ cells (EGCs) but lower levels of SOX2 and OCT4. Transfection with both SOX2 and OCT4 together was required to induce PGCs to a pluripotent state at an efficiency of 1.71%, and the further addition of C-MYC in...
Therapeutic hypothermia and temperature management, 2014
Local and general hypothermia are used to treat spinal cord injury (SCI), as well as other neurol... more Local and general hypothermia are used to treat spinal cord injury (SCI), as well as other neurological traumas. While hypothermia is known to provide significant therapeutic benefits due to its neuroprotective nature, it is unclear how the treatment may affect healthy tissues or whether it may cause undesired temperature changes in areas of the body that are not the targets of treatment. We performed 2-hour moderate general hypothermia (32°C core) or local hypothermia (30°C spinal cord) on rats that had received either a moderate contusive SCI or laminectomy (control) while monitoring temperatures at three sites: the core, spinal cord, and cortex. First, we identified that injured rats that received general hypothermia exhibited larger temperature drops at the spinal cord (-3.65°C, 95% confidence intervals [CIs] -3.72, -3.58) and cortex (-3.64°C, CIs -3.73, -3.55) than uninjured rats (spinal cord: -3.17°C, CIs -3.24, -3.10; cortex: -3.26°C, CIs -3.34, -3.17). This was found due to ...
Conference proceedings : ... Annual International Conference of the IEEE Engineering in Medicine and Biology Society. IEEE Engineering in Medicine and Biology Society. Annual Conference, 2013
Hypothermia has been shown to be an effective treatment for spinal cord injury. Local hypothermia... more Hypothermia has been shown to be an effective treatment for spinal cord injury. Local hypothermia is advantageous because it avoids inducing systemic side effects of general hypothermia while providing the opportunity for greater temperature reduction at the site of injury, which may contribute to increased neuroprotection. We report a new semi-invasive method for inducing local hypothermia in rats' spinal cords. Our method does not require laminectomy or penetration of the dura and is more effective at cooling the cord than transcutaneous approaches. We show that we were successfully able to cool the spinal cord to 30.2 ± 0.3°C for 2 hours with rectal temperature maintained at 37.3 ± 0.3°C after a spinal cord contusion injury. We also validated our method in control rats that received only a laminectomy. Furthermore, this method was able to reliably cool and rewarm the cord at a steady rate (Δ5.5°C in 30 min, or 0.2°C/min). Future work will include validating long-term function...
Conference proceedings : ... Annual International Conference of the IEEE Engineering in Medicine and Biology Society. IEEE Engineering in Medicine and Biology Society. Annual Conference, 2011
Spinal cord injury (SCI) causes a number of physiological and neurological changes resulting in l... more Spinal cord injury (SCI) causes a number of physiological and neurological changes resulting in loss of sensorimotor function. Recent work has shown that the central nervous system is capable of plastic behaviors post-injury, including axonal regrowth and cortical remapping. Functional integrity of afferent sensory pathways can be quantified using cortical somatosensory evoked potentials (SSEPs) recorded upon peripheral limb stimulation. We implanted 15 rats with transcranial screw electrodes and recorded SSEPs from cortical regions corresponding to each limb before and after a mild or moderate contusion injury. We report a post-injury increase in the mean amplitude of cortical SSEPs upon forelimb stimulation. SSEP amplitudes for mild and moderate SCI groups increased by 183% ± 95% and 107% ± 38% over baseline, respectively, while hindlimb SSEPs decreased by 58% ± 14% and 79% ± 4%. In addition, we report increased SSEP amplitude measured from the anatomically adjacent hindlimb regio...
Conference proceedings : ... Annual International Conference of the IEEE Engineering in Medicine and Biology Society. IEEE Engineering in Medicine and Biology Society. Annual Conference, 2012
We describe the feasibility of using diffusion tensor magnetic resonance imaging (DT-MRI) to stud... more We describe the feasibility of using diffusion tensor magnetic resonance imaging (DT-MRI) to study a contusive model of rat spinal cord injury following human stem cell transplantation at and around the site of injury. Rats receiving either a laminectomy or contusion injury were transplanted with oligodendrocyte precursor cells (OPCs). During the course of the study, bioluminescence imaging (BLI; up to 100 days) and somatosensory evoked potentials (SSEPs; up to 42 days) were used to evaluate cell survival and functional outcomes. Spinal cords were then analyzed ex vivo upon termination using diffusion tensor imaging (DTI). Improvements in fractional anisotropy (FA) at day 100 post-transplantation corresponded with cell survival and functional SSEP improvements. Thus, we illustrate the feasibility of DTI for evaluating axonal integrity in SCI after cell replacement therapies, and we provide examples utilizing OPC transplantations in a contusion rat model.
Induced pluripotent stem (iPS) cells are at the forefront of research in regenerative medicine an... more Induced pluripotent stem (iPS) cells are at the forefront of research in regenerative medicine and are envisaged as a source for personalized tissue repair and cell replacement therapy. Here, we demonstrate for the first time that oligodendrocyte progenitors (OPs) can be derived from iPS cells generated using either an episomal, non-integrating plasmid approach or standard integrating retroviruses that survive and differentiate into mature oligodendrocytes after early transplantation into the injured spinal cord. The efficiency of OP differentiation in all 3 lines tested ranged from 40% to 60% of total cells, comparable to those derived from human embryonic stem cells. iPS cell lines derived using episomal vectors or retroviruses generated a similar number of early neural progenitors and glial progenitors while the episomal plasmid-derived iPS line generated more OPs expressing late markers O1 and RIP. Moreover, we discovered that iPS-derived OPs (iPS-OPs) engrafted 24 hours followi...
Object Unilateral contusions represent an increasingly popular model for studying the pathways an... more Object Unilateral contusions represent an increasingly popular model for studying the pathways and recovery mechanisms of spinal cord injury (SCI). Current studies rely heavily on motor behavior scoring and histological evidence to make assessments. Electrophysiology represents one way to reliably quantify the functionality of motor pathways. The authors sought to quantify the functional integrity of the bilateral motor and sensory pathways following unilateral SCI by using measurements of motor and somatosensory evoked potentials (MEPs and SSEPs, respectively). Methods Eighteen rats were randomly divided into 3 groups receiving a mild unilateral contusion, a mild midline contusion, or a laminectomy only (control). Contusions were induced at T-8 using a MASCIS impactor. Electrophysiological analysis, motor behavior scoring, and histological quantifications were then performed to identify relationships among pathway conductivity, motor function, and tissue preservation. Results Hindl...
Primordial germ cells (PGCs) share many properties with embryonic stem cells (ESCs) and innately ... more Primordial germ cells (PGCs) share many properties with embryonic stem cells (ESCs) and innately express several key pluripotency-controlling factors, including OCT4, NANOG, and LIN28. Therefore, PGCs may provide a simple and efficient model for studying somatic cell reprogramming to induced pluripotent stem cells (iPSCs), especially in determining the regulatory mechanisms that fundamentally define pluripotency. Here, we report a novel model of PGC reprogramming to generate iPSCs via transfection with SOX2 and OCT4 using integrative lentiviral. We also show the feasibility of using non-integrative approaches for generating iPSC from PGCs using only these two factors. We show that human PGCs express endogenous levels of KLF4 and C-MYC protein at levels similar to embryonic germ cells (EGCs) but lower levels of SOX2 and OCT4. Transfection with both SOX2 and OCT4 together was required to induce PGCs to a pluripotent state at an efficiency of 1.71%, and the further addition of C-MYC in...
Therapeutic hypothermia and temperature management, 2014
Local and general hypothermia are used to treat spinal cord injury (SCI), as well as other neurol... more Local and general hypothermia are used to treat spinal cord injury (SCI), as well as other neurological traumas. While hypothermia is known to provide significant therapeutic benefits due to its neuroprotective nature, it is unclear how the treatment may affect healthy tissues or whether it may cause undesired temperature changes in areas of the body that are not the targets of treatment. We performed 2-hour moderate general hypothermia (32°C core) or local hypothermia (30°C spinal cord) on rats that had received either a moderate contusive SCI or laminectomy (control) while monitoring temperatures at three sites: the core, spinal cord, and cortex. First, we identified that injured rats that received general hypothermia exhibited larger temperature drops at the spinal cord (-3.65°C, 95% confidence intervals [CIs] -3.72, -3.58) and cortex (-3.64°C, CIs -3.73, -3.55) than uninjured rats (spinal cord: -3.17°C, CIs -3.24, -3.10; cortex: -3.26°C, CIs -3.34, -3.17). This was found due to ...
Conference proceedings : ... Annual International Conference of the IEEE Engineering in Medicine and Biology Society. IEEE Engineering in Medicine and Biology Society. Annual Conference, 2013
Hypothermia has been shown to be an effective treatment for spinal cord injury. Local hypothermia... more Hypothermia has been shown to be an effective treatment for spinal cord injury. Local hypothermia is advantageous because it avoids inducing systemic side effects of general hypothermia while providing the opportunity for greater temperature reduction at the site of injury, which may contribute to increased neuroprotection. We report a new semi-invasive method for inducing local hypothermia in rats' spinal cords. Our method does not require laminectomy or penetration of the dura and is more effective at cooling the cord than transcutaneous approaches. We show that we were successfully able to cool the spinal cord to 30.2 ± 0.3°C for 2 hours with rectal temperature maintained at 37.3 ± 0.3°C after a spinal cord contusion injury. We also validated our method in control rats that received only a laminectomy. Furthermore, this method was able to reliably cool and rewarm the cord at a steady rate (Δ5.5°C in 30 min, or 0.2°C/min). Future work will include validating long-term function...
Conference proceedings : ... Annual International Conference of the IEEE Engineering in Medicine and Biology Society. IEEE Engineering in Medicine and Biology Society. Annual Conference, 2011
Spinal cord injury (SCI) causes a number of physiological and neurological changes resulting in l... more Spinal cord injury (SCI) causes a number of physiological and neurological changes resulting in loss of sensorimotor function. Recent work has shown that the central nervous system is capable of plastic behaviors post-injury, including axonal regrowth and cortical remapping. Functional integrity of afferent sensory pathways can be quantified using cortical somatosensory evoked potentials (SSEPs) recorded upon peripheral limb stimulation. We implanted 15 rats with transcranial screw electrodes and recorded SSEPs from cortical regions corresponding to each limb before and after a mild or moderate contusion injury. We report a post-injury increase in the mean amplitude of cortical SSEPs upon forelimb stimulation. SSEP amplitudes for mild and moderate SCI groups increased by 183% ± 95% and 107% ± 38% over baseline, respectively, while hindlimb SSEPs decreased by 58% ± 14% and 79% ± 4%. In addition, we report increased SSEP amplitude measured from the anatomically adjacent hindlimb regio...
Conference proceedings : ... Annual International Conference of the IEEE Engineering in Medicine and Biology Society. IEEE Engineering in Medicine and Biology Society. Annual Conference, 2012
We describe the feasibility of using diffusion tensor magnetic resonance imaging (DT-MRI) to stud... more We describe the feasibility of using diffusion tensor magnetic resonance imaging (DT-MRI) to study a contusive model of rat spinal cord injury following human stem cell transplantation at and around the site of injury. Rats receiving either a laminectomy or contusion injury were transplanted with oligodendrocyte precursor cells (OPCs). During the course of the study, bioluminescence imaging (BLI; up to 100 days) and somatosensory evoked potentials (SSEPs; up to 42 days) were used to evaluate cell survival and functional outcomes. Spinal cords were then analyzed ex vivo upon termination using diffusion tensor imaging (DTI). Improvements in fractional anisotropy (FA) at day 100 post-transplantation corresponded with cell survival and functional SSEP improvements. Thus, we illustrate the feasibility of DTI for evaluating axonal integrity in SCI after cell replacement therapies, and we provide examples utilizing OPC transplantations in a contusion rat model.
Induced pluripotent stem (iPS) cells are at the forefront of research in regenerative medicine an... more Induced pluripotent stem (iPS) cells are at the forefront of research in regenerative medicine and are envisaged as a source for personalized tissue repair and cell replacement therapy. Here, we demonstrate for the first time that oligodendrocyte progenitors (OPs) can be derived from iPS cells generated using either an episomal, non-integrating plasmid approach or standard integrating retroviruses that survive and differentiate into mature oligodendrocytes after early transplantation into the injured spinal cord. The efficiency of OP differentiation in all 3 lines tested ranged from 40% to 60% of total cells, comparable to those derived from human embryonic stem cells. iPS cell lines derived using episomal vectors or retroviruses generated a similar number of early neural progenitors and glial progenitors while the episomal plasmid-derived iPS line generated more OPs expressing late markers O1 and RIP. Moreover, we discovered that iPS-derived OPs (iPS-OPs) engrafted 24 hours followi...
Object Unilateral contusions represent an increasingly popular model for studying the pathways an... more Object Unilateral contusions represent an increasingly popular model for studying the pathways and recovery mechanisms of spinal cord injury (SCI). Current studies rely heavily on motor behavior scoring and histological evidence to make assessments. Electrophysiology represents one way to reliably quantify the functionality of motor pathways. The authors sought to quantify the functional integrity of the bilateral motor and sensory pathways following unilateral SCI by using measurements of motor and somatosensory evoked potentials (MEPs and SSEPs, respectively). Methods Eighteen rats were randomly divided into 3 groups receiving a mild unilateral contusion, a mild midline contusion, or a laminectomy only (control). Contusions were induced at T-8 using a MASCIS impactor. Electrophysiological analysis, motor behavior scoring, and histological quantifications were then performed to identify relationships among pathway conductivity, motor function, and tissue preservation. Results Hindl...
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