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Biomedical subjects

Raphael C Lee

Publications and source records attributed to Raphael C Lee.

At least 19 recordsLinked to original sources
PubMed2006

Tc-99m pyrophosphate imaging of poloxamer-treated electroporated skeletal muscle in an in vivo rat model.

OBJECTIVE: This study investigates whether (99m)Tc pyrophosphate (PYP) imaging provides a quantitative non-invasive assessment of the extent of electroporation injury, and of the effect of poloxamer in vivo on electroporated skeletal muscle. METHODS: High-voltage electrical shock was used to produce electroporation injury in an anesthetized rat's hind limb. In each experiment, the injured limb was treated intravenously by either poloxamer-188, dextran, or saline, and subsequently imaged with (99m)Tc PYP. The radiotracer's temporal behavior among the experimental groups was compared using curve fitting of time-activity curves from the dynamic image data. RESULTS: The washout kinetics of (99m)Tc PYP changed in proportion to the electric current magnitude that produced electroporation. Also, (99m)Tc PYP washout from electroporated muscle differed between poloxamer-188 treatment and saline treatment. Finally, 10-kDa dextran treatment of electroporated muscle altered (99m)Tc PYP washout less than poloxamer-188 treatment. CONCLUSIONS: Behavior of (99m)Tc PYP in electroporated muscle appears to be an indicator of the amount of electroporation injury. Compared to saline, intravenous polaxamer-188 treatment reduced the amount of (99m)Tc PYP uptake. Coupled to results showing poloxamer-188 seals ruptured cellular membranes, lessens the extent of electroporation injury and improves cell viability, (99m)Tc PYP imaging appears to be a useful in vivo monitoring tool for the extent of electroporation injury.

Animals↗
PubMed2006

Surfactant copolymers prevent aggregation of heat denatured lysozyme.

We investigated the ability of certain triblock copolymer surfactant poloxamers of the form polyethylene oxide-polypropylene oxide-polyethylene oxide (PEO-PPO-PEO), to prevent formation of stable aggregates of heat denatured hen egg lysozyme. Differential scanning calorimetry (DSC) and synchrotron small angle x-ray scattering (SAXS) experiments were performed to study the thermodynamics and solution structures of lysozyme at temperatures between 20 and 90 degrees C in the presence and absence of poloxamers with various molecular weights (8.4-14.3 kDa), but similar hydrophile/hydrophobe (PEO:PPO) ratio of 80%. Poloxmer 188 was found to be very effective in preventing aggregation of heat denatured lysozyme and those functioned as a synthetic surfactant, thus enabling them to refold when the conditions become optimal. For comparison, we measured the ability of 8 kDa polyethylene glycol (PEG) to prevent lysozyme aggregation under same conditions. The results of these studies suggest that poloxamers are more efficient than PEG in preventing aggregation of heat denaturated lysozyme. To achieve equivalence, more than an order of magnitude higher concentration of PEG concentration was needed. Apparently, the presence of a hydrophobic segment in the poloxamers increases their ability to target the hydrophobic region of the unfolded proteins and protect them from self association. Given their biocompatibility and the low concentrations at which they effectively facilitate refolding of denatured proteins, they may be useful in the treatment of burns and other conditions resulting in the denaturation of proteins.

Hydrophobic and Hydrophilic Interactions↗
PubMed2006

Neuropsychological changes following electrical injury.

The clinical presentation of electrical injury commonly involves physical, cognitive, and emotional complaints. Neuropsychological studies, including case reports, have indicated that electrical injury (EI) survivors may experience a broad range of impaired neuropsychological functions, although this has not been clarified through controlled investigation. In this study, we describe the neuropsychological test findings in a series of 29 EI patients carefully screened and matched to a group of 29 demographically similar healthy electricians. Participants were matched by their estimated premorbid intellectual ability. Multivariate analysis of variance was used to assess group differences in the following neuropsychological domains: attention and mental speed, working memory, verbal memory, visual memory, and motor skills. EI patients performed significantly worse on composite measures of attention/mental speed and motor skills, which could not be explained by demographic differences, injury parameters, litigation status, or mood disturbance. Results suggest that cognitive changes do occur in patients suffering from electrical injury.

Adult↗
PubMed2005

Supramembrane potential-induced electroconformational changes in sodium channel proteins: a potential mechanism involved in electric injury.

Effects of imposed large supraphysiological transmembrane potential (TP) pulses on channel proteins, particularly on the voltage-gated Na channels, were investigated. Voltage clamp techniques were used to deliver both shock and stimulation pulses, and to monitor changes in the channel functions. Our experimental results indicated that more than one 4 ms duration TP shock of -450 mV resulted in electroconformational denature of voltage-gated Na channels. This resulted in functional reductions in muscle cells' excitability. We quantified the TP shock-induced decrease in the Na channel currents, compared the pre- and post-shocked Na channel currents' voltage dependency, and studied the reversibility of the electroconformationally denatured ion channel proteins. These observations are particularly relevant to the problem of explaining the neuromuscular damage following high voltage electrical shock injuries despite no evidence of a thermal injury component.

Animals↗
PubMed2005

Effects of crowding on the thermal stability of heterogeneous protein solutions.

Crowding can substantially affect the transition of a protein between its native (N) and unfolded (U) states via volume exclusion effects. Also, it influences considerably the aggregation (A) of unfolded proteins. To examine the details, we developed an approach for computing the kinetic rates of the process N <--> U --> A in which the concentration of the protein is explicitly taken into account. We then compute the relative change with temperature of the protein denaturation for various fractional volume occupancies and partition of proteins in solution. The analysis indicates that, in protein solutions in which the average distance between proteins is comparable with the radius of gyration of an unfolded protein, steric effects increase the stability of the proteins which are in compact, native states. In heterogeneous protein solutions containing various types of proteins with different thermal stabilities, the unfolding of the most thermolabile proteins will increase the stability of the other proteins. The results shed light on the way proteins change the thermal stability of a cell as they unfold and aggregate. This study may be valuable in questions related to the dynamics of thermal injuries.

Animals↗
PubMed2005

Molecular crowding effects on protein stability.

The volume fraction occupied by the dry matter of the cell can be as large as 40%, of which more than half (approximately 60%) are proteins. Thus, cellular proteins and protein assemblies occupy a large volume that can have a profound effect on their own native-state stabilities and on their unfolding/refolding rates. In addition, macromolecular crowding can change the properties of a significant fraction of the water in the cell. We review features of the molecular crowding effect which are relevant for describing the microscopic mechanism of thermal injuries.

Mathematics↗
PubMed2005

Cell injury by electric forces.

The molecular architecture of biological systems is heavily influenced by the highly polar interactions of water. Thus, macromolecules such as proteins that are highly water soluble must be electrically polar. Energy generation methods needed to support cell metabolic processes depend on compartmentalizing mobile ions and thus require electrical ion transport barriers such as membranes. One consequence of these biological design constraints is vulnerability to injury by electrical forces. Supraphysiological electric forces cause damage to cells and tissues by disrupting cell membranes and altering the conformation of biomolecules. In addition, prolonged passage of electrical current leads to damage by thermal mechanisms. This review will focus on the non-thermal effects.

Cell Membrane↗
PubMed2005

Multimodal strategies for resuscitating injured cells.

Our cells and tissues are challenged constantly by exposure to extreme conditions that cause acute and chronic stress. Wounding at the cellular level is a common event, and results from cell exposure to supra-physiologic forces, or is the consequence of action by reactive chemical agents. An individual cellular wound results from either the alteration of protein or DNA structure, or the disruption of molecular assemblies, the most important of which is the cell's membranes. Tissue healing at the macroscopic level is a complex and coordinated process involving many different cell types while, in contrast, the wounds of individual cells heal primarily via biomolecular interactions. Like tissue wound healing, cellular wound healing involves the upregulation or acceleration of processes that are constitutively expressed in routine physiologic repair of cellular structures In addition, recent advances have been made in the identification of pharmaceutical strategies to aid the cellular repair response. Many of these strategies offer promise for augmenting the already present cellular repair mechanisms.

Cell Membrane↗
PubMed2005

A surfactant copolymer facilitates functional recovery of heat-denatured lysozyme.

The triblock copolymer poloxamer 188 is a non-cytotoxic, nonionic surfactant with both hydrophobic and hydrophilic domains. We show that P188 is able to facilitate the recovery of catalytic activity of heat-denatured lysozyme in dilute solution at low molar ratios of P188:enzyme. Heat-denatured enzyme retained 55% of native activity. After treatment with P188, the enzyme's activity was 85% of native. Because of the low molar ratios used and the non-cytotoxic nature of the compound, P188 may be of potential use in burn therapy.

Animals↗
PubMed2005

Incidence of hypertrophic scars among African Americans linked to vitamin D-3 metabolism?

The pathogenesis and progression of wound-healing involve intricate pathways and numerous chemical mediators. This remains an area of intense study as undesirable results of this process, such as hypertrophic scars and keloids, can result in significant morbidity. These lesions are distinct in their characteristics, although they are similar in their distribution in patients with darker skin colors. There is a robust inflammatory mechanism behind the formation of hypertrophic scars and keloids. Furthermore, their development may be intimately related to vitamin D-3, which has been shown to be a powerful anti-inflammatory agent. This chemical is made in the skin, whose production is influenced by various factors of which the amount of melanin is a crucial one. More specifically, an increase in pigmentation has been shown to decrease the amount of vitamin D-3 synthesis in the skin. Thus, this paper proposes the hypothesis linking the propensity of inflammation and subsequent scarring in darker-skinned individuals to the reduced levels of vitamin D-3 production in their skin.

Black or African American↗
PubMed2004

Subcutaneous tri-block copolymer produces recovery from spinal cord injury.

We have studied the ability of nonionic detergents and hydrophilic polymers to seal permeabilized membranes of damaged cells, rescuing them from progressive dissolution, degeneration, and death. We report that a single subcutaneous injection of the tri-block copolymer, Poloxamer 188 (P188) 6 hr after a severe compression of the adult guinea pig spinal cord is able to: (1). preserve the anatomic integrity of the cord; (2). produce a rapid recovery of nerve impulse conduction through the lesion; and (3). produce a behavioral recovery of a spinal cord dependent long tract spinal cord reflex. These observations stood out against a control group in blinded evaluation. Conduction through the lesion was monitored by stimulating the tibial nerve of the hind limb, and measuring the arrival of evoked potentials at the contralateral sensory cortex of the brain (somatosensory evoked potentials; SSEP). Behavioral recovery was determined by a return of sensitivity of formerly areflexic receptive fields of the cutaneous trunchi muscle (CTM) reflex. This contraction of back skin in response to tactile stimulation is totally dependent on the integrity of an identified bilateral column of ascending long tract axons. A statistically significant recovery of both SSEP conduction through the lesion and the CTM reflex occurred in P188-treated animals compared to vehicle-treated controls. Quantitative 3D computer reconstruction of the lesioned vertebral segment of spinal cord revealed a statistically significant sparing of spinal cord parenchyma and a significant reduction in cavitation of the spinal cord compared to control animals We determined that the proportion of P188-treated animals that recovered evoked potentials were nearly identical to that produced by a subcutaneous injection of polyethylene glycol (PEG). In contrast, P188 was not as effective as PEG in producing a recovery of CTM functioning. We discuss the likely differences in the mechanisms of action of these two polymers, and the possibilities inherent in a combined treatment.

Animals↗
PubMed2004

Electroporation of a lipid bilayer as a chemical reaction.

When a cell's transmembrane potential is increased from a physiological one to more than 370 mV, the transmembrane current increases more than hundredfold within a millisecond. This is due to the formation of conductive pores in the membrane. We construct a model in which we conceive of pore formation as a voltage sensitive chemical reaction. The model predicts the logarithm of the pore formation rate to increase proportionally to the square of the voltage. We measure currents through frog muscle cell membranes under 8 ms pulses of up to 440 mV. The experimental data appear consistent with the model.

Animals↗
PubMed2004

Poloxamer 188 prevents acute necrosis of adult skeletal muscle cells following high-dose irradiation.

Acute cellular necrosis occurring minutes to hours after massive ionizing radiation exposure (IR) results from rapid membrane lipid peroxidation, blebbing and membrane breakdown. We have shown, previously, that certain polymer surfactants can restore structural integrity and transport barrier function of cell membranes following high-dose IR. We now investigate, specifically, the efficacy of the amphiphilic surfactant Poloxamer 188 (P188) in preventing acute necrosis of adult rat skeletal muscle cells after high-dose IR. Explanted cells were treated with 60Co IR doses of 10, 40 or 80Gy and their viability was determined using fluorometric probes at 4 and 18h post-IR. IR of 10Gy did not cause acute necrosis. Significant acute cell necrosis was observed after 40 and 80Gy doses in a dose-dependent manner. Post-IR treatment with P188 significantly enhanced the cells' viability post-IR treatment. By comparison 10kDa neutral dextran, a hydrophilic polymer, was found to be ineffective. Despite progressive cell death over 18h after high-dose IR, cells treated with P188 manifested greater survival than media or dextran-treated cells. It appears that use of P188 or similar multi-block copolymers to prolong viability of irradiated cells in vitro through membrane sealing is an important step in development of effective interventional therapy for extreme IR exposure. Not only can repairing the membrane prevent acute necrosis, but it also can provide a critical time opportunity to address other mechanisms of cell death, such as apoptosis or mitotic arrest, which manifest over a longer time frame.

Animals↗
PubMed2004

Rectovaginal radiation fistula repair using an obturator fasciocutaneous thigh flap.

BACKGROUND: Rectovaginal fistulae are a known complication of pelvic radiotherapy utilizing locally applied isotope implants. Most often, either permanent colostomy or reconstruction with a well-vascularized flap is necessary. Traditional techniques for fistula repair utilize bulky muscle flaps, disfiguring pudendal artery flaps or may require laparotomy. CASE: We describe the management of a 26-year-old woman with a large radiation-induced rectovaginal fistula. A fasciocutaneous medial thigh flap based on terminal branches of the obturator artery and vein was used without colostomy and resulted in pain-free sexual function and minimal vulva disfigurement. CONCLUSION: A medial thigh fasciocutaneous flap without muscle can be transferred into the vagina on the obturator vessels and may become the preferred method for managing large rectovaginal fistulas.

Adenocarcinoma↗
PubMed2004

Poloxamer 188 volumetrically decreases neuronal loss in the rat in a time-dependent manner.

OBJECTIVE: Excitotoxicity is a multistep process that results in either necrosis or apoptosis. It has been associated with neuronal death in trauma, ischemia, and neurodegeneration. The final step in necrotic cell death is the rupture of a cell's plasma membrane; repair of this membrane rupture is a potentially powerful technique of neuroprotection. Poloxamer 188 (P-188) is a synthetic surfactant that seals experimentally porated membranes. This study investigated the usefulness and time dependence of intrathecal P-188 in protecting neurons in an in vivo model of excitotoxicity in the rat. METHODS: Twenty-eight Sprague-Dawley rats underwent striatal infusion of quinolinic acid to produce a spherical excitotoxic lesion. Each animal then received either vehicle or P-188 at 10 minutes, 4 hours, or both time points after surgery by direct cisterna magna injection. Animals were killed at 1 week, and brains were stained immunohistochemically for the neuronal marker Neu-N. Volumes of neuronal loss were calculated and compared between groups by analysis of variance. RESULTS: All animals were found to have spherical, stereotyped lesions. The animals that received intrathecal poloxamer at the early injection time had statistically smaller lesions (8.16 +/- 6.12 mm(3); n = 5; P = 0.0015) than controls (18.25 +/- 11.42 mm(3); n = 11). Those animals that received poloxamer at both injection times also had statistically smaller lesions (10.57 +/- 9.00 mm(3); n = 7; P = 0.0095). The group that received poloxamer at the late injection time only did not have significantly decreased lesion size (14.86 +/- 7.95 mm(3); n = 5). CONCLUSION: Intrathecal P-188 reduces neuronal loss after excitotoxic injury in the rat only when delivered immediately after the toxin. This observation confirms the potential of surfactant molecules as neuroprotectants but predicts that their usefulness is best realized by early and potentially ongoing treatment.

Animals↗
PubMed2004

Surfactant poloxamer 188-related decreases in inflammation and tissue damage after experimental brain injury in rats.

OBJECT: The surfactant, poloxamer 188 (P-188), has been found to protect against tissue injury in various experimental models. Its protective mechanism may involve the effects of the surfactant against oxidative stress and inflammation. The authors investigated the role of P- 188 in the reduction of tissue injury and macrophage response in a model of excitotoxic brain injury in the rat striatum. METHODS: Fifteen Sprague-Dawley rats underwent stereotactic injection of 120 nmol of quinolinic acid into the striatum and received intracisternal injection of vehicle or P-188 (40 mg/kg) at 10 minutes and 4 hours postinjury. Rats were killed after 1 week, and the histological score was determined based on the degree of overall tissue injury (Grades 1-4) at the lesion site. The number of macrophages within the lesioned striatum was compared with that found within the striatum on the nonoperated contralateral side. The scores related to tissue damage and the macrophage ratios of each group were then compared using t-tests. Striatal injection of the toxin produced a lesion characterized by necrosis and inflammation surrounding the injection site in all six control animals. In rats in which intracisternal P-188 was administered, significantly less tissue injury was demonstrated (mean score 2.45 +/- 0.74) than in controls (mean score 3.14 +/- 0.75) (p = 0.045). The rats that received intracisternal surfactant also had significantly less macrophage infiltrate (mean ratio 1.06 +/- 0.18) than control animals (mean ratio 2.00 +/- 0.48) (p = 0.004). CONCLUSIONS: The surfactant P-188 reduces tissue loss and macrophage infiltrate after excitotoxic brain injury in the rat. Possible mechanisms of this effect may include direct surfactant modulation of inflammatory cell membrane fluidity.

Animals↗
PubMed2004

Prevention and treatment of excessive dermal scarring.

Today, wound management to avoid excessive scar formation is increasingly important, especially in populations with Fitzpatrick 3 or higher skin pigmentation. Medical science and industrial development are devoting more effort toward understanding and offering better therapy to control scars. However, advances in scar management have been hampered by the confusing or ambiguous terminology. There is no consensus on what amount of post-traumatic skin scar formation is "normal" and what should be considered "hypertrophic". In the World Health Organization's ICD-9, there is no diagnostic code for hypertrophic scar--only keloid is listed. Yet, the medical and scientific literature distinguishes them as different conditions. Our experience suggests that the diagnosis of keloid disease is greatly over-rendered. For black patients, an elevated scar seems, by default, diagnosed as keloid by most. This confusion results in inappropriate management of scar formation, and occasionally contributes to decision making related to elective or cosmetic surgery. Given that patients are expecting better outcomes from wound care today than in the past, this review article attempts to capture the essential biological factors related to wound scar production and discusses treatment options and indications used by the authors.

Anti-Inflammatory Agents↗