Abstract WMP42: Astrocyte Specific rAAV-mediated Insulin-Like Growth Factor-1 Expression in Aging Female Rats Increases Post-Stroke Survival and Sensory Motor Performance

Background and Purpose: Our previous work shows that middle aged female rats sustain larger strokes as compared to younger female rats. With age, circulating and brain parenchymal levels of IGF-1 are reduced. Exogenous IGF-1 treatment improves infarct volume in aging females. Our recent studies show that astrocytes from aging females synthesize less IGF-1. Here we tested the hypothesis that elevation of astrocyte derived IGF-1 would improve stroke impairment in older female rats. Methods: Middle-aged (10-12 month old; acyclic) female rats were injected with adeno-associated virus serotype 5 (rAAV5) into the cortex and striatum. rAAV5 was packaged with the coding sequence of the IGF-1 gene downstream of an astrocyte-specific gene (GFAP). The construct contained the mCherry reporter gene. Control rAAV consisted of an identical shuttle vector construct without the IGF-1 gene. In separate experiments, two titers of virus were injected: high dose (5 X 10 12 VP/mL) or low dose (5 X 10 11 VP/mL). Three to four weeks after injection, middle-cerebral artery occlusion via an intraluminal suture for ninety minutes was performed followed by reperfusion. Post-surgery survival was monitored as well as sensory motor function using the vibrissae evoked forelimb placement task. Results: Specificity of IGF-1 expression was confirmed by visualization of the mCherry reporter under fluorescent illumination and immunohistochemistry. Post stroke survival was improved in animals that received the high dose rAAV-IGF-1 animals a 5-day period (p<0.001). Low dose rAAV-IGF-1 did not affect post stroke survival, however sensory motor performance was preserved in this group. In low dose control animals, ischemic stroke impaired performance on the vibrissae evoked forelimb placement task. Impairment was seen in the same-side and cross-midline task performance on the limb contralateral to the infarct and cross midline task on the limb ipsilateral to the infarcted side (p<0.05). No significant deficits were seen in the rAAV-IGF-1 low dose treated animals. Conclusion: These data support the hypothesis that increasing astrocytic IGF-1 in aging females improves post stroke survival and behavior outcomes.

Download Full-text

Abstract WP96: The Histone Deacetylase Inhibitor, Sodium Butyrate, Exhibits Biphasic Neuroprotective Mechanisms in the Acute Phase of Ischemic Stroke in Middle-Aged Female Rats

Stroke ◽

10.1161/str.48.suppl_1.wp96 ◽

2017 ◽

Vol 48 (suppl_1) ◽

Author(s):

Min Jung Park ◽

Farida Sohrabji

Keyword(s):

Histone Deacetylase ◽

Acute Phase ◽

Sodium Butyrate ◽

Hdac Inhibitor ◽

Female Rats ◽

Middle Aged ◽

Peripheral Tissues ◽

Post Stroke ◽

Sensory Motor ◽

Anti Inflammatory

Introduction: Sodium butyrate (NaB) is a histone deacetylase (HDAC) inhibitor exhibiting anti-inflammatory and neuroprotective effects in a rat ischemic model of a myocardial ischemia as well as stroke. Although clinical evidence shows that older women are at higher risk for stroke occurrence and greater stroke severity, no studies have evaluated the effectiveness of NaB either in females or in older animals. Methods: To determine the effects of NaB on stroke in older females, acyclic middle-aged Sprague-Dawley female rats (10-12 months old, constant diestrus) were subject to middle cerebral artery occlusion (MCAo) by intracerebral injection of recombinant endothelin-1. Rats were treated with NaB (300 mg/kg, i.p.) at 6h and 30h following ET-1 injection. Animals were tested for sensory motor performance pre and post stroke. Subsequently, rats were sacrificed at the early (2d) or late (5d) acute phase after MCAo. Serum and tissue samples were collected for biochemical analyses. Results: NaB treatment reduced infarct volume and ameliorated stroke-induced sensory motor impairment in middle-aged female rats post MCAo. At the early acute phase, NaB treatment decreased brain lipid peroxides, and reduced serum levels of GFAP, a marker of blood brain barrier permeability. NaB also reduced expression of the inflammatory cytokine IL-1beta in circulation and IL-18 in the ischemic hemisphere. At the late acute phase, NaB treatment further suppressed MCAo-induced increase of IL-1beta, IL-17A, and IL-18 in brain lysates (cortex and striatum) from the ischemic hemisphere, and decreased ischemia-induced upregulation of IL-1beta and IL-18 in circulation, indicating a potent anti-inflammatory effect of the HDAC inhibitor. Moreover, NaB treatment also increased expression of IGF-1, a known neuroprotectant, in peripheral tissues including serum, liver, and spleen. Conclusions: These data provide the first evidence that delayed (> 6h) NaB treatment post-stroke is neuroprotective in older female rats. Importantly, these data also show that in addition to its well-known anti-inflammatory actions, NaB may exert a biphasic effect after stroke, operating initially to reduce oxidative stress in the brain, and later, elevating IGF-1 expression in peripheral tissues.

Download Full-text

Abstract WP108: Astrocyte-specific rAAV-mediated Insulin-like Growth Factor-1 Expression in Aging Female Rats Alters Stroke-induced Brain Immune Profile and Reduces Motor Impairment

Stroke ◽

10.1161/str.48.suppl_1.wp108 ◽

2017 ◽

Vol 48 (suppl_1) ◽

Author(s):

Andre K Okoreeh ◽

Shameena Bake ◽

Farida Sohrabji

Keyword(s):

Flow Cytometry ◽

Growth Factor ◽

Immune Cells ◽

Motor Impairment ◽

Female Rats ◽

Insulin Like Growth Factor ◽

Middle Aged ◽

Post Stroke ◽

Sensory Motor ◽

The Brain

Introduction: Middle-aged female rats sustain larger strokes than younger female rats. This may be due to age-related loss of circulating and parenchymal brain levels of insulin like growth factor-1 (IGF-1). With age, IGF-1 synthesis is decreased in astrocytes, a critical cell type for post-stroke recovery. Here we tested the hypothesis that replenishing IGF-1 in aging astrocytes will improve stroke outcomes by influencing the type and extent of immune cells infiltration into the brain post-stroke. Methods: Middle-aged (10-12 mo old, acyclic) female rats were injected into the striatum and cortex with adeno-associated virus serotype 5 (rAAV5) packaged with the coding sequence of the hIGF-1 gene downstream of an astrocyte-specific promoter (GFAP). The rAAV-control consisted of an identical shuttle vector construct without the hIGF-1 gene. Three to four weeks later, animals underwent 90 minute transient middle cerebral artery occlusion via intraluminal suture. At 2d post stroke, flow cytometry was used to determine the type and extent of peripheral immune cells trafficked into the brain. In parallel studies, animals were tested for performance on sensory motor tasks at 2d and 5 days after MCAo. Results: rAAV-mediated IGF-1 expression was confirmed in astrocytes with RT-PCR. Flow cytometry analysis of immune cells in the brain at 24h post-stroke found that proportion of Treg cells was greater in animals with rAAV-IGF-1 as compared to rAAV-controls. Additionally, while there was no difference in the proportion of M2 microglia, rAAV-IGF-1 enhanced M2 infiltrating macrophages. At 2d and 5d post stroke, stroke-induced sensory motor impairment was reduced in animals with rAAV-IGF-1 as compared to rAAV-controls. Conclusion: Targeted enhancement of IGF-1 in astrocytes of middle-aged females improved stroke-induced behavioral impairment concomitant with recruitment of anti-inflammatory cell types to the ischemic brain.

Download Full-text

Functional Assessment of Stroke-Induced Regulation of miR-20a-3p and Its Role as a Neuroprotectant

Translational Stroke Research ◽

10.1007/s12975-021-00945-x ◽

2021 ◽

Author(s):

Taylor E. Branyan ◽

Amutha Selvamani ◽

Min Jung Park ◽

Kriti E. Korula ◽

Kelby F. Kosel ◽

...

Keyword(s):

Motor Behavior ◽

Mitochondrial Dynamics ◽

Infarct Volume ◽

Neuron Specific Enolase ◽

Cell Types ◽

Female Rats ◽

Therapeutic Window ◽

Middle Aged ◽

Stroke Outcomes ◽

Sensory Motor

AbstractMicroRNAs have gained popularity as a potential treatment for many diseases, including stroke. This study identifies and characterizes a specific member of the miR-17–92 cluster, miR-20a-3p, as a possible stroke therapeutic. A comprehensive microRNA screening showed that miR-20a-3p was significantly upregulated in astrocytes of adult female rats, which typically have better stroke outcomes, while it was profoundly downregulated in astrocytes of middle-aged females and adult and middle-aged males, groups that typically have more severe stroke outcomes. Assays using primary human astrocytes and neurons show that miR-20a-3p treatment alters mitochondrial dynamics in both cell types. To assess whether stroke outcomes could be improved by elevating astrocytic miR-20a-3p, we created a tetracycline (Tet)-induced recombinant adeno-associated virus (rAAV) construct where miR-20a-3p was located downstream a glial fibrillary acidic protein promoter. Treatment with doxycycline induced miR-20-3p expression in astrocytes, reducing mortality and modestly improving sensory motor behavior. A second Tet-induced rAAV construct was created in which miR-20a-3p was located downstream of a neuron-specific enolase (NSE) promoter. These experiments demonstrate that neuronal expression of miR-20a-3p is vastly more neuroprotective than astrocytic expression, with animals receiving the miR-20a-3p vector showing reduced infarction and sensory motor improvement. Intravenous injections, which are a therapeutically tractable treatment route, with miR-20a-3p mimic 4 h after middle cerebral artery occlusion (MCAo) significantly improved stroke outcomes including infarct volume and sensory motor performance. Improvement was not observed when miR-20a-3p was given immediately or 24 h after MCAo, identifying a unique delayed therapeutic window. Overall, this study identifies a novel neuroprotective microRNA and characterizes several key pathways by which it can improve stroke outcomes.

Download Full-text

Male gender increases sensitivity to renal injury in response to cholesterol loading

AJP Renal Physiology ◽

10.1152/ajprenal.00009.2002 ◽

2003 ◽

Vol 284 (4) ◽

pp. F718-F726 ◽

Cited By ~ 17

Author(s):

Diana M. Attia ◽

Roel Goldschmeding ◽

Mahmoud A. Attia ◽

Peter Boer ◽

Hein A. Koomans ◽

...

Keyword(s):

Renal Injury ◽

Low Dose ◽

Plasma Cholesterol ◽

A Priori ◽

No Synthase ◽

Male Gender ◽

Female Rats ◽

Male Rats ◽

High Dose ◽

Cholesterol Feeding

Males are at greater risk for renal injury than females. This may relate to nitric oxide (NO) availability, because female rats have higher renal endothelial NO synthase (NOS) levels. Previously, our laboratory found susceptibility to proteinuria induced by NOS inhibition in male compared with female rats. Dyslipidemia and hypercholesterolemia dose dependently decreased renal NOS activity and caused renal injury in female rats. We hypothesized that exposure of male rats to hypercholesterolemia would lead to more renal injury in male than in female rats due to an a priori lower renal NO system. Female and male rats were fed no, low-dose, or high-dose cholesterol for 24 wk. Cholesterol feeding dose dependently increased proteinuria in both female and male rats, but male rats developed more proteinuria at similar plasma cholesterol ( P < 0.001). Control males had lower renal NOS activity than control females (4.44 ± 0.18 vs. 7.46 ± 0.37 pmol · min−1 · mg protein−1; P < 0.05), and cholesterol feeding decreased renal NOS activity in males and in females ( P < 0.05). Cholesterol-fed males developed significantly more vascular, glomerular, and tubulointerstitial monocyte/macrophage influx and injury than females. Thus under baseline conditions, male rats have lower renal NOS activity than female rats. This may explain why male rats are more sensitive to renal injury by factors that decrease NO availability, such as hypercholesterolemia.

Download Full-text

Abstract TP104: Post Stroke Depressive-Like Behaviors in Middle-Aged Female Rats is Associated With Gut Dysbiosis and Improved by Mir363-3p

Stroke ◽

10.1161/str.51.suppl_1.tp104 ◽

2020 ◽

Vol 51 (Suppl_1) ◽

Author(s):

Courtney Stewart ◽

Aditya Panta ◽

Taylor Branyan ◽

Farida Sohrabji

Keyword(s):

Cell Proliferation ◽

Dentate Gyrus ◽

Forced Swim Test ◽

Female Rats ◽

Middle Aged ◽

16S Sequencing ◽

Post Stroke ◽

Hippocampal Cell ◽

Affective Behaviors ◽

Post Stroke Depression

Background: Post-stroke depression (PSD) occurs in one-third of stroke survivors, is more prevalent in women, and negatively impacts quality of life and recovery. Our previous work showed that acyclic middle-aged female rats display depressive behaviors after stroke, and IV treatment with mir363-3p attenuates these behaviors. Serotonin has been implicated in affective behaviors, and loss of this neurotransmitter is also associated with decreased neurogenesis in the hippocampus. The present study determined (a) whether stroke affected tryptophan, a gut metabolite, and precursor for serotonin, and cell proliferation in the hippocampus and (b) whether serum tryptophan levels and hippocampal cell proliferation was restored by mir363-3p. Methods: Ischemic stroke was induced by stereotaxic delivery of endothelin-1 to the MCA of middle-aged female rats. Animals received a single injection of mir363-3p or scrambled oligos 4h after stroke. Depressive-like behaviors were assessed by the Effort-based sucrose consumption test, Social Interaction and Forced Swim Test 3 months after stroke. Blood was collected at termination. Serum tryptophan levels were quantified by ELISA. Bacterial composition was analyzed by 16S sequencing of fecal samples. Ki67 immunohistochemistry was performed on brain tissue collected at termination and quantified within the dentate gyrus by two blind observers. Results: Animals subjected to stroke exhibited depressive-like behaviors and had decreased tryptophan levels as compared to sham treated animals. This is also accompanied by microbiota dysbiosis as measured by an elevated ratio of Firmicutes to Bacteroidetes. Mir363-3p treatment increased tryptophan levels as compared to scrambled controls and were no different from sham (non-stroke) animals. Moreover, miR363-3p treated rats exhibit a significantly more dentate gyrus-specific Ki67 expression a proliferation marker. Conclusions: Together, these data indicate that miR363-3p attenuates post stroke depression via a mechanism that prevents microbiota dysbiosis and increases hippocampal cell proliferation.

Download Full-text

Efficacy of a low-dose melatonin pretreatment in protecting against the neurobehavioral consequences of chronic hypoperfusion in middle-aged female rats

Behavioural Brain Research ◽

10.1016/j.bbr.2019.112257 ◽

2020 ◽

Vol 377 ◽

pp. 112257 ◽

Cited By ~ 1

Author(s):

Maxine De Butte ◽

Blake Gieseking

Keyword(s):

Low Dose ◽

Female Rats ◽

Middle Aged

Download Full-text

Unrestricted Hepatocyte Transduction with Adeno-Associated Virus Serotype 8 Vectors in Mice

Journal of Virology ◽

10.1128/jvi.79.1.214-224.2005 ◽

2005 ◽

Vol 79 (1) ◽

pp. 214-224 ◽

Cited By ~ 221

Author(s):

Hiroyuki Nakai ◽

Sally Fuess ◽

Theresa A. Storm ◽

Shin-ichi Muramatsu ◽

Yuko Nara ◽

...

Keyword(s):

Portal Vein ◽

Dose Response ◽

Smooth Muscles ◽

The Body ◽

Specific Gene ◽

High Dose ◽

Adeno Associated Virus ◽

Virus Serotype ◽

High Doses ◽

Dose Response Study

ABSTRACT Recombinant adeno-associated virus (rAAV) vectors can mediate long-term stable transduction in various target tissues. However, with rAAV serotype 2 (rAAV2) vectors, liver transduction is confined to only a small portion of hepatocytes even after administration of extremely high vector doses. In order to investigate whether rAAV vectors of other serotypes exhibit similar restricted liver transduction, we performed a dose-response study by injecting mice with β-galactosidase-expressing rAAV1 and rAAV8 vectors via the portal vein. The rAAV1 vector showed a blunted dose-response similar to that of rAAV2 at high doses, while the rAAV8 vector dose-response remained unchanged at any dose and ultimately could transduce all the hepatocytes at a dose of 7.2 × 1012 vector genomes/mouse without toxicity. This indicates that all hepatocytes have the ability to process incoming single-stranded vector genomes into duplex DNA. A single tail vein injection of the rAAV8 vector was as efficient as portal vein injection at any dose. In addition, intravascular administration of the rAAV8 vector at a high dose transduced all the skeletal muscles throughout the body, including the diaphragm, the entire cardiac muscle, and substantial numbers of cells in the pancreas, smooth muscles, and brain. Thus, rAAV8 is a robust vector for gene transfer to the liver and provides a promising research tool for delivering genes to various target organs. In addition, the rAAV8 vector may offer a potential therapeutic agent for various diseases affecting nonhepatic tissues, but great caution is required for vector spillover and tight control of tissue-specific gene expression.

Download Full-text

Carcinogenesis Bioassay of Propyl Gallate in F344 Rats and B6C3FJ Mice

Journal of the American College of Toxicology ◽

10.3109/10915818309140729 ◽

1983 ◽

Vol 2 (6) ◽

pp. 425-433 ◽

Cited By ~ 12

Author(s):

K. M. Abdo ◽

J. E. Huff ◽

J. K. Haseman ◽

M. P. Dieter ◽

G. A. Boorman ◽

...

Keyword(s):

Propyl Gallate ◽

Dose Group ◽

Low Dose ◽

Female Rats ◽

Male Rats ◽

High Dose Group ◽

High Dose ◽

Preputial Gland ◽

Rats And Mice ◽

F344 Rats

Chronic toxicity studies were conducted by maintaining groups of 50 F344 rats and 50 B6C3F1 mice of each sex on nutritionally complete diets containing 0%, 0.6%, or 1.2% propyl gallate for 103 weeks. Survival of rats and mice of both sexes was not significantly affected by the administration of this compound. Dosed rats and mice showed growth retardation and reduced feed utilization efficiency. Increased incidence of hepatic cytoplasmic vacuolization and suppurative inflammation of the prostate gland were observed in dosed male rats and were considered to be related to propyl gallate administration. Tumors of the preputial gland, islet ceil tumors of the pancreas, and pheochromocytoma of the adrenal gland were observed with significantly (p < 0.05) higher incidence in the low-dose male rats; however, there was little evidence of a dose response or of an effect in the high-dose group. Rare tumors (an astrocytoma and a glioma) were found in the brains of two low-dose female rats but none was found in the high-dose group. Malignant lymphoma occurred with a significant (p < 0.05) positive trend in male mice and the incidence in the high-dose group was significantly (p < 0.05) higher than that of the concurrent controls. However, the high-dose incidence was not significantly different from the historical control rate for the laboratory that conducted the bioassay. Under the conditions of the bioassay, propyl gallate was not considered to be clearly carcinogenic for F344 rats, although the increased incidence of preputial gland tumors, islet-cell tumors of the pancreas, and pheochromocytoma of the adrenal glands in low-dose male rats may have been related to compound administration. Thus, the evidence for carcinogenicity in male rats is regarded as being equivocal, while there was no indication of a carcinogenic response in female rats. Propyl gallate was not considered to be carcinogenic for B6C3F1 mice, although the increased incidence of malignant lymphoma in dosed male mice may have been related to administration of the test compound.

Download Full-text

Subchronic Toxicological Assessment of Dr Iguedo Goko Cleanser® on Lipid Profile and Serum Antioxidant Enzymes in Exposed Wistar Rats

Asian Journal of Biology ◽

10.9734/ajob/2020/v9i430093 ◽

2020 ◽

pp. 16-25

Author(s):

Godswill J. Udom ◽

Jude E. Okokon ◽

John A. Udobang ◽

Daniel N. Obot ◽

Nkechi J. Onyeukwu

Keyword(s):

Oxidative Stress ◽

Lipid Profile ◽

Wistar Rats ◽

Low Dose ◽

Herbal Remedy ◽

Female Rats ◽

Male Rats ◽

High Density Lipoproteins ◽

High Dose ◽

Low Density

Dr Iguedo Goko Cleanser® is a polyherbal mixture promoted as an effective herbal remedy for numerous diseases. Study aimed to evaluate the toxicity concern of the polyherbal mixture (PHM) on lipid profile and oxidative status in Wistar rats of both gender. Acute toxicity study was conducted using modified method of Lorke. Thirty Wistar rats of bother gender were randomly divided into six groups (5/group) and exposed to the polyherbal mixture for 60 days via oral gavage. Control groups (1 and 4) received 10 mL/kg distilled water, while groups 2-3 and 5-6 received 476.24 and 158.75 mg/kg body weight of Dr Iguedo Goko Cleanser® respectively. On 62nd day, animals were sacrificed under diethyl ether anaesthesia; blood samples were collected by cardiac puncture for biochemical analysis. PHM significantly (p < 0.05) increased high density lipoproteins (HDL) levels in male rats as well as high dose female rats relative to control. However, low dose female rats recorded low HDL levels relative to control. Total cholesterol, triglycerides, low density and very low density lipoprotein levels were significantly reduced in all test groups relative to controls. The low dose males (LDM) had reduced serum glutathione peroxidase (GPX) activity; while increased and decreased GPX and glutathione (GSH) activities were respectively recorded for female rats. Male rats had dose-dependent increase in malondialdehyde. The recorded reductions in serum lipids suggest that the polyherbal mixture may have hypolipidemic potentials. While the increased malondialdehyde as well as decreased GPX and GSH indicate lipid peroxidation and oxidative stress inducing potentials of the PHM. Despite the positive modulation on lipid profile, findings suggest utmost caution on chronic use as its oxidative stress inducing potentials is considerable.

Download Full-text