Biomedical subjects

J W Simpkins

Publications and source records attributed to J W Simpkins.

At least 19 recordsLinked to original sources

PubMed2001

Synthesis and biological evaluation of 17beta-alkoxyestra-1,3, 5(10)-trienes as potential neuroprotectants against oxidative stress.

17beta-O-Alkyl ethers (methyl, ethyl, propyl, butyl, hexyl, and octyl) of estradiol were obtained from 3-O-benzyl-17beta-estradiol with sodium hydride/alkyl halide, followed by the removal of the O-benzyl protecting group via catalytic transfer hydrogenation. An increase compared to estradiol in the protection of neural (HT-22) cells against oxidative stress due to exposure of glutamate was furnished by higher (C-3 to C-8) alkyl ethers, while methyl and ethyl ethers decreased the neuroprotective effect significantly. Lipophilic (butyl and octyl) ethers blocking the phenolic hydroxyl (3-OH) of A-ring were inactive.

Animals↗

PubMed2001

Membrane fluidity effects of estratrienes.

Estrogens have demonstrable neuroprotective effects. This fact has lead to the proposed use of estrogens for the prevention and/or treatment of Alzheimer's disease. The exact protective mechanism estrogens provide is not fully understood. In this report, a potential non-genomic mechanism for estratrienes involving alterations in membrane fluidity was studied. Steroids, such as estrogen, are known to be membrane-active and can alter the lipid packing. In this study we used fluorescent methodologies to address the effect of naturally occurring steroids (17alpha and 17beta-estradiol, testosterone, and progesterone) and new estratriene analogs on membrane fluidity using liposomes and HT-22 hippocampal cells. The study's results indicate steroids, based on the estratriene nucleus, can modulate lipid packing as evidenced by (1) decreased membrane fusion events and (2) decreased membrane fluidity. The effects on the membrane were both time and concentration dependent. It was also demonstrated through rational design estratriene analogs can be synthesized with enhanced membrane effects. Finally, in a glutamate-induced toxicity HT-22 model, we also demonstrated cellular protection with the estratriene-based molecules and analogs. The data suggest the plethora of cellular actions of estrogens may relate to or be influenced by membrane effects of the steroid.

Alzheimer Disease↗

PubMed2001

Estradiol protects against ATP depletion, mitochondrial membrane potential decline and the generation of reactive oxygen species induced by 3-nitroproprionic acid in SK-N-SH human neuroblastoma cells.

Mitochondria are recognized as modulators of neuronal viability during ischemia, hypoxia and toxic chemical exposure, wherein mitochondria dysfunction leading to ATP depletion may be a common pathway of cell death. Estrogens have been reported to be neuroprotective and proposed to play a role in the modulation of cerebral energy/glucose metabolism. To address the involvement of 17beta-estradiol preservation of mitochondrial function, we examined various markers of mitochondrial activity in human SK-N-SH neuroblastoma cells exposed to 3-nitroproprionic acid (3-NPA), a succinate dehydrogenase inhibitor which uncouples oxidative phosphorylation. 3-NPA (10 mM) significantly increased ATP levels at 2 h then caused a 40% and a 50% decrease in ATP levels from baseline when treated for 12 h and 24 h, respectively. 3-NPA also induced significant increases in levels of cellular hydrogen peroxide and peroxynitrite at 2 h and a 60% decrease in mitochondrial membrane potential (MMP) at 12 h exposure. 17beta-Estradiol (17beta-E(2)) pretreatment restored the ATP level back to 80% at 12 h of that in control cells treated with 3-NPA but without E(2), blunted the effect of 3-NPA on MMP and reactive oxygen species levels. The present study indicates that 17beta-E(2) can preserve mitochondrial function in the face of inhibition of oxidative phosphorylation.

Adenosine Triphosphate↗

PubMed2001

17-beta estradiol can reduce secondary ischemic damage and mortality of subarachnoid hemorrhage.

Subarachnoid hemorrhage (SAH) is a unique disorder commonly occurring when an aneurysm ruptures, leading to bleeding and clot formation, with a higher incidence in females. To evaluate the influence of 17-beta estradiol (E2) in the outcome of subarachnoid hemorrhage, SAH was induced by endovascular puncture of the intracranial segment of internal carotid artery in 15 intact females (INT), 19 ovariectomized females (OVX), and 13 ovariectomized female rats with E2 replacement (OVX + E2). Cerebral blood flow was recorded before and after SAH. All animals were decapitated immediately after death or 24 hours after SAH for clot area analysis. Brains were sliced and stained with 2,3,5-triphenyltetrazolium chloride (TTC) for secondary ischemic lesion analysis. The cortical cerebral blood flow (CBF), which was measured by a laser-Doppler flowmeter, decreased to 29.6%+/-17.7%, 22.8%+/-8.3%, and 43.5%+/-22.9% on the ipsilateral side (P = 0.01), and decreased to 63.4%+/-14.1%, 57.4%+/-11.0%, and 66.6%+/-17.9% on the contralateral side (P = 0.26) in INT, OVX, and OVX + E2, respectively. The subcortical CBF, which were measured by the H2 clearance method, were 7.77+/-12.03, 7.80+/-8.65, and 20.58+/-8.96 mL 100 g(-1) min(-1) on the ipsilateral side (P < 0.01), and 21.53+/-2.94, 25.13+/-3.01, and 25.30+/-3.23 mL 100 g(-1) min(-1) on the contralateral side in INT, OVX, and OVX + E2, respectively. The mortality was 53.3%, 68.4%, and 15.4% in INT, OVX, and OVX + E2, respectively (P = 0.01), whereas no significant difference in clot area was noted among the groups. The secondary ischemic lesion volume was 9.3%+/-8.4%, 24.3%+/-16.3%. and 7.0%+/-6.4% in INT, OVX, and OVX + E2, respectively (P < 0.01). This study demonstrated that E2 can reduce the mortality and secondary ischemic damage in a SAH model without affecting the clot volume.

Animals↗

PubMed2001

Estrogens decrease reperfusion-associated cortical ischemic damage: an MRI analysis in a transient focal ischemia model.

BACKGROUND AND PURPOSE: Early identification of irreversible cerebral ischemia is critical in defining strategies that influence neuronal survival after stroke. We used MRI to investigate the effects of 17beta-estradiol (E2) on the temporal evolution of focal ischemia. METHODS: Female rats were ovariectomized and divided into 1 of 2 groups: ovariectomy alone (OVX; n=4) or ovariectomy with estrogen replacement (OVX+E2; n=3). Both groups were then subjected to 1-hour middle cerebral artery occlusion (MCAO), with the use of a standardized endovascular monofilament model, followed by reperfusion. Sequential diffusion-weighted (DWI) and T2-weighted (T2WI) MRI were obtained during and after the MCAO. In separate groups of animals (n=5 for OVX and OVX+E2), cerebral blood flow (CBF) was measured by laser-Doppler methods before, during, and after occlusion. RESULTS: DWI detected similar lesion characteristics during MCAO in both groups. In the OVX group, lesion size did not change during reperfusion, but the signal intensity ratio increased early and stabilized during the latter stages. In contrast, DWI lesion size decreased during reperfusion in OVX+E2 rats by 50% to 60% (P<0.05), a size reduction almost exclusively limited to cortical regions. During MCAO, the signal intensity ratio in OVX+E2 rats was reduced compared with OVX rats. Reperfusion further attenuated the signal intensity ratio in cortical but not subcortical regions (P<0.05 versus OVX). T2WI revealed no lesions in either group during MCAO, but it detected lesion sizes similar to that of DWI during reperfusion. Furthermore, similar patterns and magnitudes of estrogen treatment-related decrease in lesion size were noted after reperfusion. T2WI demonstrated less intense signal intensity ratio changes in both groups compared with DWI. There were no differences in CBF between groups either during occlusion, early reperfusion, or 1 day after reperfusion. CONCLUSIONS: This study strongly suggests that estrogens selectively protect cortical tissue from ischemic damage during MCAO and that this protection is exerted during both the occlusion and reperfusion phases of ischemia and does not involve an estrogen-related change in CBF.

Animals↗

PubMed2001

The nonfeminizing enantiomer of 17beta-estradiol exerts protective effects in neuronal cultures and a rat model of cerebral ischemia.

Estrogens are potent neuroprotective compounds in a variety of animal and cell culture models, and data indicate that estrogen receptor (ER)-mediated gene transcription is not required for some of these effects. To further address the requirement for an ER in estrogen enhancement of neuronal survival, we assessed the enantiomer of 17beta-estradiol (ENT-E(2)), which has identical chemical properties but interacts only weakly with known ERs, for neuroprotective efficacy. ENT-E(2) was both as potent and efficacious as 17beta-estradiol in attenuating oxidative stress-induced death in HT-22 cells, a murine hippocampal cell line. Further, ENT-E(2) completely attenuated H(2)O(2) toxicity in human SK-N-SH neuroblastoma cells at a 10 nM concentration. In a rodent model of focal ischemia, 17beta-estradiol (100 microgram/kg) or ENT-E(2) (100 microgram/kg), injected 2 h before middle cerebral artery occlusion, resulted in a 60 and 61% reduction in lesion volume, respectively. ENT-E(2), at the doses effective in this study, did not stimulate uterine growth or vaginal opening in juvenile female rats when administered daily for 3 days. These data indicate that the neuroprotective effects of estrogens, both in vitro and in vivo, can be disassociated from the peripheral estrogenic actions.

Animals↗

PubMed2000

Definition of the anterior choroidal artery territory in rats using intraluminal occluding technique.

This manuscript delineates the territory of the anterior choroidal artery (AChA) in rats, as defined by the induction of an AChA infarction. By advancing a 0.24-mm surgical suture up the internal carotid artery (ICA) to a point 0.5-2 mm proximal to the middle cerebral artery (MCA) origin, the AChA could be occluded and a reliable AChA distribution infarction was produced in 62% (23/37) of animals. The infarct volume, as defined by TTC staining, was 55+/-7 mm(3). Maps of the infarction, generated by measuring the entire area of overlapping coronal slices, demonstrated that the internal capsule was always damaged. Other areas that might be affected included the hippocampus, thalamus, amygdaloid complex, piriform cortex, dorsal caudatoputamen, and lateral ventricular wall. Positioning the coated suture proximal to the AChA produced a much smaller infarct involving the medial and lateral hypothalamus, preoptic region, optic chiasm, and marginal region of the internal capsule near to the lateral hypothalamus exempt from AChA territory damage. A causative relationship between AChA occlusion and a deep cerebral infarct centered on the internal capsule was further established by: (1) identifying the AChA on the non-ischemic side with colored silicone perfusion, and subsequent similar delineation on the ischemic side, and (2) delineating infarction in the silicone perfused AChA region using hematoxylin and eosin staining and the TUNEL method. The AChA usually originated from the ICA (91% of cases), 1.75+/-0.12 mm proximal to the MCA bifurcation. Approximately 27% of the AChAs had periamygdaloid branch(es) on its initial segment.

Animals↗

PubMed2000

Hypoperfusion induces overexpression of beta-amyloid precursor protein mRNA in a focal ischemic rodent model.

Silent stroke is one of the risk factors of dementia. In the present study, we used a novel focal ischemic animal model to investigate the effects of comparatively small changes of cerebral blood flow (CBF) on the expression of beta-amyloid precursor protein (APP) mRNA. Focal ischemia was achieved by introducing a 4-0 monofilament to the bifurcation of anterior and middle cerebral arteries. Brain samples were harvested from ischemic core and penumbra of cortices at 1, 4 and 7 days following ischemia. The expression of APP mRNA was assessed by RT-PCR. The CBF was decreased to 50% for 1 day after stroke and recovered to 90% at the fourth day after stroke. The changes of CBF were accompanied by an increase in the expression of APP mRNA. APP mRNA increased to 208% and 152% in the penumbra and core ischemic regions, respectively, on the fourth day after MCAO and remained high through the seventh day of ischemia. This study suggests brain hypoperfusion enhances APP mRNA expression and may contribute to the progression of cognitive impairment after silent stroke.

Amyloid beta-Protein Precursor↗

PubMed2000

Neuroprotective effects of phenolic A ring oestrogens.

We have formulated the 'phenolic A ring hypothesis' for the neuroprotective effects of oestrogens based upon several observations: (i) structure-activity relationships show that a phenolic A ring and at least two additional rings are required for neuroprotection while oestrogenicity requirements are more stringent; (ii) neuroprotection with phenolic A ring compounds occurs in cells that lack oestrogen receptors and are not antagonized by anti-oestrogens; (iii) phenolic A ring compounds rapidly activate a variety of signal transduction pathways that are known to be involved in cell homeostasis; and (iv) in vivo, treatment with oestrogens results in a neuronal type-independent neuronal protection from ischaemic insult. Potential mechanisms of actions that may be involved in the neuroprotective effects of phenolic A ring compounds are: (i) oestrogen redox cycling that potently inhibits oxidative stress; (ii) interactions with signal transduction pathways including the transcription factor cAMP response element binding protein; and (iii) induction of anti-apoptotic proteins. These signalling pathways may individually or collectively contribute to the plethora of neuronal cell types that are protected from a variety of insults by oestrogen-like compounds.

Animals↗

PubMed2000

Protective effect of estrogens against oxidative damage to heart and skeletal muscle in vivo and in vitro.

Estrogen has been shown to protect skeletal muscle from damage and to exert antioxidant properties. The purpose of the present study was to investigate the antioxidant and protective properties of estrogens in rodent cardiac and skeletal muscle and H9c2 cells. Female Sprague-Dawley rats were separated into three groups, ovariectomized (OVX), ovariectomized with estrogen replacement (OVX + E2), and intact control (SHAM), and were assessed at two time periods, 4 and 8 weeks. Rodents hearts were analyzed for basal and iron-stimulated lipid peroxidation in the absence and presence of beta-estradiol (betaE2) by measuring thiobarbituric acid reactive species (TBARS). Isolated soleus (SOL) and extensor digitorum longus (EDL) were analyzed for creatine kinase (CK) efflux. Using H9c2 cells, the in vitro effects of betaE2 and its isomer alpha-estradiol were investigated under glucose-free/hypoxic conditions. TBARS assay was also performed on the H9c2 in the presence or absence of betaE2. The results indicate that OVX rodent hearts are more susceptible to lipid peroxidation than OVX + E2 hearts. OVX soleus showed higher cumulative efflux of CK than OVX + E2. Furthermore, H9c2 survival during oxidative stress was enhanced when estrogen was present, and both OVX hearts at 4 weeks and H9c2 cells particularly were protected from oxidative damage by estrogens. We conclude that estrogen protects both skeletal and cardiac muscle from damage, and its antioxidant activity can contribute to this protection.

Animals↗

PubMed2000

Estrogens and estrogen-like non-feminizing compounds. Their role in the prevention and treatment of Alzheimer's disease.

The present position paper is intended to provide evidence that estrogen deprivation contributes to the occurrence and course of Alzheimer's disease (AD) and that currently available estrogen preparations may be useful in the prevention and treatment of AD in women. Additionally, there is now substantial preclinical evidence to support the development of novel non-feminizing estrogens for use in male and female subjects for the protection of neurons from damage and death that underlies the neuropathology of AD. Estrogens and non-feminizing estrogen-like compounds may exert their beneficial effects in AD through a variety of mechanisms, directly through their neuroprotective actions and indirectly through their neurotrophic effects. Inasmuch as estrogens are comparatively free of both acute and chronic toxicities, and non-feminizing estrogens are expected to be even safer, their use for years to decades for the prevention or treatment of AD is possible.

Alzheimer Disease↗

PubMed2000

Estradiol exerts neuroprotective effects when administered after ischemic insult.

BACKGROUND AND PURPOSE: 17beta-Estradiol (E2) has been reported to exert neuroprotective effects when administered before an ischemic insult. This study was designed to determine whether E2 treatment after ischemia exerts the same effects and, if so, how long this therapeutic window remains open, and whether the effects are related to changes in cerebral blood flow (CBF). METHODS: Female Sprague-Dawley rats were subjected to permanent middle cerebral artery occlusion (MCAO). In protocol 1, E2 was administered (100 microg/kg IV followed immediately by subcutaneous implantation of crystalline E2 in a silicone elastomer tube) to ovariectomized females (OVX+E2) at 0.5 (n=8), 1 (n=6), 2 (n=7), 3 (n=6), or 4 (n=9) hours after MCAO. Intact (INT; n=6) and ovariectomized females (OVX; n=12) were subjected to MCAO and received vehicle instead of E2. Two days after MCAO the animals were killed, and ischemic lesion volume was determined by 2,3,5-triphenyltetrazolium chloride staining. In protocol 2, CBF was monitored before and at 1, 24, and 48 hours in a group of animals receiving E2 or vehicle 0.5 hour after ischemia induction (INT, n=6; OVX, n=8; OVX+E2, n=6). RESULTS: Lesion volume was 20.9+/-2.2% and 21.8+/-1.2% in the INT and OVX groups, respectively. E2 was found to decrease lesion volume significantly when administered within 3 hours after MCAO. The lesion volumes were 6.3+/-0.5%, 10.3+/-2.1%, 11.8+/-1.8%, 13.5+/-1.6%, and 17.9+/-2.8% when E2 was administered at 0.5, 1, 2, 3, or 4 hours after MCAO, respectively. CBF decreased to 43.1+/-2.2% and 25.4+/-1.0% in the INT and OVX animals, respectively, at 5 minutes after MCAO. In comparison to OVX rats, CBF was not different at 1 hour after E2 administration but was increased significantly in the OVX+E2 group 1 and 2 days after E2 administration. CONCLUSIONS: E2 exerts neuroprotective effects when administered after ischemia, with a therapeutic window in a permanent focal cerebral ischemia model of approximately 3 hours. This effect of estradiol was associated with no immediate change in blood flow but with a delayed increase in CBF.

Animals↗

PubMed1999

The mammary tumor response in triazine-treated female rats: a threshold-mediated interaction with strain and species-specific reproductive senescence.

Triazine herbicides are among the most heavily used agricultural pesticides. Although they possess a very low acute toxicity in animals, a mammary tumor response has been consistently observed in Sprague-Dawley (SD) female rats following chronic oral dosing of atrazine and simazine at and above maximum tolerated doses. However, a substantial collection of detailed research has clearly shown that triazines are not genotoxic or mutagenic, nor do they possess estrogenic agonist activity that might promote mammary tumor growth. Examination of estrous cycling records of atrazine-treated SD rats revealed a premature appearance of persistent estrous episodes, beyond the prevalent occurrence normally seen in untreated, aging SD rats. A significant correlation has been found between early or severe estrous cycle disruption of atrazine-treated rats and the early appearance of mammary tumors. In studies using SD female rats fed atrazine for 6 months, then ovariectomized and administered an estrogen-containing silastic s.c. implant, a deficient luteinizing hormone surge was observed at a 400 parts per million (ppm) dose, but not at 25 or 50 ppm. Because SD rats exhibiting persistent estrus also have a prolonged elevation of estrogen secretion, it is proposed that the triazine-associated mammary tumor response is promoted by the test animal's own estrogen from ovarian follicles that fail to ovulate because gonadotropin surge sufficiency is blocked by the high dose of herbicide. It is further proposed that, because reproductive senescence in SD rats is fundamentally different from menopause in women, the animal response to dosing, as well as the enormous requisite dosing level, establishes a safety margin of very low risk to human health from this mode of action.

Aging↗

PubMed1999

Glutamate receptor requirement for neuronal death from anoxia-reoxygenation: an in Vitro model for assessment of the neuroprotective effects of estrogens.

1. Previous studies demonstrated that estrogens, specifically 17 beta-estradiol, the potent, naturally occurring estrogen, are neuroprotective in a variety of models including glutamate toxicity. The aim of the present study is twofold: (1) to assess the requirement for glutamate receptors in neuronal cell death associated with anoxia-reoxygenation in three cell types, SK-N-SH and HT-22 neuronal cell lines and primary rat cortical neuronal cultures, and (2) to evaluate the neuroprotective activity of both 17 beta-estradiol and its weaker isomer, 17 alpha-estradiol, in both anoxia-reoxygenation and glutamate toxicity. 2. SK-N-SH and HT-22 cell lines, both of which lack NMDA receptors as assessed by MK-801 binding assays, were resistant to both anoxia-reoxygenation and glutamate-induced cell death. In contrast, primary rat cortical neurons, which exhibit both NMDA and AMPA receptors, were sensitive to brief periods of exposure to anoxia-reoxygenation or glutamate. As such, there appears to be an obligatory requirement for NMDA and/or AMPA receptors in neuronal cell death resulting from brief periods of anoxia followed by reoxygenation. 3. Using primary rat cortical neuronal cultures, we evaluated the neuroprotective activity of 17 beta-estradiol (1.3 or 133 nM) and 17 alpha-estradiol (133 nM) in both anoxia-reoxygenation and excitotoxicity models of cell death. We found that the 133 nM but not the 1.3 nM dose of the potent estrogen, 17 beta-estradiol, protected 58.0, 57.5, and 85.3% of the primary rat cortical neurons from anoxia-reoxygenation, glutamate, or AMPA toxicity, respectively, and the 133 nM dose of the weak estrogen, 17 alpha-estradiol, protected 74.6, 81.7, and 85.8% of cells from anoxia-reoxygenation, glutamate, or AMPA toxicity, respectively. These data demonstrate that pretreatment with estrogens can attenuate glutamate excitotoxicity and that this protection is independent of the ability of the steroid to bind the estrogen receptor.

2-Hydroxypropyl-beta-cyclodextrin↗

PubMed1999

Experimental model of small deep infarcts involving the hypothalamus in rats: changes in body temperature and postural reflex.

BACKGROUND AND PURPOSE: Intraluminal middle cerebral artery (MCA) occlusion in rats has been reported to cause hyperthermia assumed to be caused by hypothalamic damage. To clarify the effects of hypothalamic ischemia on body temperature and to obtain a model simulating lacunar infarction, we attempted to produce small infarcts in deep structures (including the hypothalamus). METHODS: A surgical suture was advanced to occlude the origin of the hypothalamic (HTA) and/or anterior choroidal arteries (AChA) without compromise of the anterior or middle cerebral artery origins. After treatment, rectal temperature and postural reflex were examined repeatedly for 3 days under nonanesthetic conditions. The AChA and HTA and their link with small deep infarction were then confirmed by TTC, hematoxylin and eosin, and TUNEL stains and by microsurgical dissection after colored silicone perfusion into the cerebral arteries. RESULTS: Advancement of the suture near to but not occluding the MCA origin (0.5 to 1.9 mm proximal) produced small, deep, nonneocortical strokes in 25 of 36 animals without producing MCA ischemic changes. These infarctions mainly affected the hypothalamus in 13 animals (HTA area: infarct volume 6+/-1 mm(3)) and involved both the internal capsule and hypothalamus in 12 animals (HTA+AChA area infarct volume 48+/-10 mm(3)). Rats with HTA infarction alone exhibited persistent hyperthermia for 72 hours; some also had transient mild postural abnormality. The AChA+HTA infarct group showed a transient elevation of body temperature for 24 hours and definitive postural abnormality. In the remaining 11 animals, the suture was inadvertently advanced across the MCA origin, producing a large infarct that affected both the neocortex (MCA territory) and nonneocortical structures (volume 381+/-30 mm(3), n=11). The MCA infarct group displayed a transient hyperthermia and severe postural abnormality. CONCLUSIONS: When properly positioned, the intraluminal suture method permits selective AChA and/or HTA obstruction without inducing MCA territory ischemia. This model confirms that selective hypothalamic infarction produces significant and sustained temperature regulation abnormalities. The model also may be useful in investigating the pathophysiology of small, deep, end-vessel infarction.

Animals↗

PubMed1999

Disposition of morphine in tissues of the pregnant rat and foetus following single and continuous intraperitoneal administration to the mother.

Foetal exposure to maternally administered opiates such as morphine represent a serious human health problem but disposition studies in man are difficult to perform. Morphine disposition was therefore investigated in pregnant rats and their foetuses near term as a model. Disposition was examined either following intraperitoneal dosing as a single dose or continuous infusion. A high-pressure liquid chromatography assay for morphine in plasma and tissue was developed and validated. Following the single morphine dose, foetal distribution was rapid and concentrations in foetal and placental tissue were from 2.6 (whole foetus) to 27.6 (placenta) times higher compared with maternal plasma. The rank order of the area under the concentration vs time curve (AUC) of morphine in tissues was: placenta > or = foetal liver > foetal brain > whole foetus > maternal brain. The foetal brain to maternal brain AUC ratio for morphine was 9.5, suggesting large differences in their blood-brain barrier permeability. Following continuous administration of morphine there were significant linear relationships between maternal plasma and tissue concentrations with the same rank order as the single dose study. However, following continuous administration the relative amount of morphine in placenta and foetal liver was reduced by half and one-third, respectively, compared with the single dose study. These results document why the rat foetus is particularly susceptible to the pharmacodynamic effects of morphine following maternal administration.

Analgesics, Opioid↗

PubMed1999

Pre-exposure of cells to cationic lipids enhances transgene delivery and expression in a tissue culture cell line.

Several factors influence non-viral transfection in tissue culture models including nature of the cationic lipid, plasmid construction, and DNA lipid complex, among others. The cell line itself is another confounding variable. Each subcellular population may respond independently to the transgene or specific delivery vector with regards to toxicity or transgene expression. In this study, the SKnSH (human neuroblastoma) and COS-1 (African green kidney) cells were exposed to three different treatments A, B, and C. Treatment A refers to cells obtained from American Type Culture Collection (ATCC) and cultivated as recommended, treatment B to cells that were grown in presence of cationic lipids for two weeks, and treatment C to cells that were grown in presence of cationic lipids for two weeks followed by normal media for two weeks to determine if lipid mediated effects were reversible. Treatment B resulted in a three-fold increase in transgene expression of a reporter gene as compared to the other treatments. This increase in transgene expression appeared not to be related to alterations in toxicity. Interestingly, the fluid phase endocytic uptake of fluorescently labeled oligonucleotides was increased in treatment B. However, there was no significant difference in the cellular-associated signal when fluorescently labeled plasmid-DNA was evaluated. In COS-1 cells, no difference in transfection was observed with treatment B illustrating that cell lines respond independently. In conclusion, pre-exposure of SKnSH cells to cationic liposomes (treatment B) resulted in higher transgene production.

Animals↗

PubMed1998

Estrogen attenuates over-expression of beta-amyloid precursor protein messager RNA in an animal model of focal ischemia.

Cerebral ischemia is a risk factor for late onset Alzheimer's disease. Since estrogen replacement therapy benefits the outcome of cerebral stroke in post-menopausal women, we designed the present study to investigate the effects of estrogen on the expression of beta-amyloid precursor protein (APP) mRNA following focal ischemia in female rats. Female rats were ovariectomized (OVX) for two weeks. A single dose of 17 beta-estradiol (E2) (100 microgram/kg) was injected s.c. two hours before a unilateral middle cerebral artery (MCA) occlusion. Brain samples were harvested from ischemic core and penumbra of cortices at one hour and twenty-four hours following MCA occlusion. The expression of APP mRNA was assessed by RT-PCR. At one hour after MCA occlusion, OVX rats had a 67.9% (p<0.05) increase in APP mRNA in the penumbra. E2 treatment reduced this APP mRNA over-expression by 26.3% at that region. At twenty four hours following MCA occlusion, OVX rats had increases in APP mRNA of 52.9% and 57.0% (p<0.05) in the core and penumbra, respectively. E2 treatment reduced the APP mRNA over-expression by 61.0% and 48.6% (p<0.05) in these two regions, respectively. These effects appeared to reflect an interaction between hormonal environment and ischemia, since in the absence of MCA occlusion, there were no significant differences in APP mRNA expression among OVX, OVX-E2 treated and intact female rats. The present study demonstrates that estrogen may have an important role in reducing the over-expression of APP mRNA following focal ischemia.

Amyloid beta-Protein Precursor↗