Microthrombi Correlates With Infarction and Delayed Neurological Deficits After Subarachnoid Hemorrhage in Mice

Background and Purpose: Delayed neurological deficits are a devastating consequence of subarachnoid hemorrhage (SAH), which affects about 30% of surviving patients. Although a very serious concern, delayed deficits are understudied in experimental SAH models; it is not known whether rodents recapitulate the delayed clinical decline seen in SAH patients. We hypothesized that mice with SAH develop delayed functional deficits and that microthrombi and infarction correlate with delayed decline. Methods: Adult C57BL/6J mice of both sexes were subjected to endovascular perforation to induce SAH. Mice were allowed to survive for up to 1 week post-ictus and behavioral performance was assessed daily. Postmortem microthrombi, large artery diameters (to assess vasospasm), and infarct volume were measured. These measures were analyzed for differences between SAH mice that developed delayed deficits and SAH mice that did not get delayed deficits. Correlation analyses were performed to identify which measures correlated with delayed neurological deficits, sex, and infarction. Results: Twenty-three percent of males and 47% of females developed delayed deficits 3 to 6 days post-SAH. Female mice subjected to SAH had a significantly higher incidence of delayed deficits than male mice with SAH. Mice that developed delayed deficits had significantly more microthrombi and larger infarct volumes than SAH mice that did not get delayed deficits. Microthrombi positively correlated with infarct volume, and both microthrombi and infarction correlated with delayed functional deficits. Vasospasm did not correlate with either infarction delayed functional deficits. Conclusions: We discovered that delayed functional deficits occur in mice following SAH. Sex differences were seen in the prevalence of delayed deficits. The mechanism by which microthrombi cause delayed deficits may be via formation of infarcts.

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Sex Differences in the Relation Between Frailty and Endothelial Dysfunction in Old Mice

The Journals of Gerontology Series A ◽

10.1093/gerona/glab317 ◽

2021 ◽

Author(s):

Jazmin A Cole ◽

Mackenzie N Kehmeier ◽

Bradley R Bedell ◽

Sahana Krishna Kumaran ◽

Grant D Henson ◽

...

Keyword(s):

Sex Differences ◽

Endothelial Function ◽

Vascular Function ◽

Mesenteric Artery ◽

Frailty Index ◽

Mesenteric Arteries ◽

Male Mice ◽

Male And Female ◽

Female Mice ◽

Age Related

Abstract Vascular endothelial function declines with age on average, but there is high variability in the magnitude of this decline within populations. Measurements of frailty, known as frailty index (FI), can be used as surrogates for biological age, but it is unknown if frailty relates to the age-related decline in vascular function. To examine this relation, we studied young (4-9 months) and old (23-32 months) C57BL6 mice of both sexes. We found that FI was greater in old compared with young mice, but did not differ between old male and female mice. Middle cerebral artery (MCA) and mesenteric artery endothelium-dependent dilation (EDD) also did not differ between old male and female mice; however, there were sex differences in the relations between FI and EDD. For the MCA, FI was inversely related to EDD among old female mice, but not old male mice. In contrast, for the mesenteric artery, FI was inversely related to EDD among old male mice, but not old female mice. A higher FI was related to a greater improvement in EDD with the superoxide scavenger TEMPOL in the MCAs for old female mice and in the mesenteric arteries for old male mice. FI related to mesenteric artery gene expression negatively for extracellular superoxide dismutase (Sod3) and positively for interleukin-1β (Il1b). In summary, we found that the relation between frailty and endothelial function is dependent on sex and the artery examined. Arterial oxidative stress and pro-inflammatory signaling are potential mediators of the relations of frailty and endothelial function.

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Abstract T P210: Splenectomy Attenuates Sex Differences in Infarct Volume Following Experimental Stroke in Mice

Stroke ◽

10.1161/str.45.suppl_1.tp210 ◽

2014 ◽

Vol 45 (suppl_1) ◽

Author(s):

Jianming Wang ◽

Sheetal Bodhankar ◽

Halina Offner ◽

Stephanie J Murphy

Keyword(s):

Sex Differences ◽

Blood Glucose ◽

Brain Injury ◽

Infarct Volume ◽

Blood Gases ◽

Experimental Stroke ◽

Ischemic Brain Injury ◽

Ischemic Brain ◽

Male And Female ◽

Female Mice

It is now increasingly clear that human stroke can have other serious consequences besides brain damage that can impact on patient survival and recovery. For example, many stroke patients succumb to CNS injury-induced immunodepression and fatal infections. Our prior work suggests that evolving cerebral ischemic injury elicits a cycle of injury from brain-to-spleen-to-brain that is strongly influenced by sex. We determined if splenic immunocytes are important in contributing to sex differences in post-ischemic brain injury. Male and female C57BL/6J mice were splenectomized 14 days before experimental stroke. Male and female mice with or without splenectomy (n=9-10 per group) then underwent 60 min of middle cerebral artery occlusion (MCAO) via intraluminal filament. Laser-Doppler flowmetry (LDF) was used to monitor cortical perfusion. All mice were euthanized and brains collected at 96 hours of reperfusion. Infarct volume (% corrected contralateral structure) was determined by image analysis of coronal brain slices stained with 2,3,5-triphenyltetrazolium chloride. Mean arterial blood pressure (MABP), blood gases (pH, P a O 2 , P a CO 2 ), and blood glucose were measured at 30 min MCAO and at 15 min of reperfusion in separate groups of male and female mice with or without splenectomy (n=5 per group). Relative LDF changes (% baseline), MABP, blood gases, and blood glucose during and after MCAO were comparable among the experimental groups. We observed that infarct volume in females (cortex, 41±4%; striatum, 55±6%) was smaller ( P <0.05) compared to males (cortex, 52±3%; striatum, 75±3%) at 96 hours of reperfusion. However, no differences (cortex, P =0.313; striatum, P =0.601) in infarct volume were seen between splenectomized male (cortex, 43±4%; striatum, 51±7%) and female (cortex, 38±4%; striatum, 46±5%) mice. Our data suggest that removal of all splenocyte lineages via splenectomy attenuates sex differences in post-ischemic brain injury. Future studies will evaluate the role of different splenic immunocyte subsets, such as T or B lymphocytes, on male vs. female ischemic brain outcomes. This study was supported by National Institutes of Health grant NS076013.

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Sex differences in renal and metabolic responses to a high-fructose diet in mice

AJP Renal Physiology ◽

10.1152/ajprenal.00403.2014 ◽

2015 ◽

Vol 308 (5) ◽

pp. F400-F410 ◽

Cited By ~ 17

Author(s):

Nikhil Sharma ◽

Lijun Li ◽

C. M. Ecelbarger

Keyword(s):

Sex Differences ◽

Glucose Transporter ◽

Collecting Duct ◽

Urine Volume ◽

Thick Ascending Limb ◽

Male Mice ◽

Male And Female ◽

Female Mice ◽

High Fructose Diet ◽

High Fructose

High fructose intake has been associated with increased incidences of renal disease and hypertension, among other pathologies. Most fructose is cleared by the portal system and metabolized in the liver; however, systemic levels of fructose can rise with increased consumption. We tested whether there were sex differences in the renal responses to a high-fructose diet in mice. Two-month-old male and female C57BL6/129/SV mice ( n = 6 mice per sex per treatment) were randomized to receive control or high-fructose (65% by weight) diets as pelleted chow ad libitum for 3 mo. Fructose feeding did not significantly affect body weight but led to a 19% and 10% increase in kidney weight in male and female mice, respectively. In male mice, fructose increased the expression (∼50%) of renal cortical proteins involved in metabolism, including glucose transporter 5 (facilitative fructose transporter), ketohexokinase, and the insulin receptor (β-subunit). Female mice had lower basal levels of glucose transporter 5, which were unresponsive to fructose. However, female mice had increased urine volume and plasma K+ and decreased plasma Na+ with fructose, whereas male mice were less affected. Likewise, female mice showed a two- to threefold reduction in the expression Na+-K+-2Cl− cotransporter 2 in the thick ascending limb and aquaporin-2 in the collecting duct with fructose relative to female control mice, whereas male mice had no change. Overall, our results support greater proximal metabolism of fructose in male animals and greater distal tubule/collecting duct (electrolyte homeostasis) alterations in female animals. These sex differences may be important determinants of the specific nature of pathologies that develop in association with high fructose consumption.

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Mechanisms Contributing to Cerebral Infarct Size after Stroke: Gender, Reperfusion, T Lymphocytes, and Nox2-Derived Superoxide

Journal of Cerebral Blood Flow & Metabolism ◽

10.1038/jcbfm.2010.14 ◽

2010 ◽

Vol 30 (7) ◽

pp. 1306-1317 ◽

Cited By ~ 109

Author(s):

Vanessa H Brait ◽

Katherine A Jackman ◽

Anna K Walduck ◽

Stavros Selemidis ◽

Henry Diep ◽

...

Keyword(s):

Ischemic Stroke ◽

T Lymphocytes ◽

Infarct Volume ◽

Ischemia Reperfusion ◽

Fold Increase ◽

Cerebral Infarct ◽

Male Mice ◽

Female Mice ◽

Underlying Mechanisms ◽

Cerebral Reperfusion

Cerebral infarct volume is typically smaller in premenopausal females than in age-matched males after ischemic stroke, but the underlying mechanisms are poorly understood. In this study we provide evidence in mice that this gender difference only occurs when the ischemic brain is reperfused. The limited tissue salvage achieved by reperfusion in male mice is associated with increased expression of proinflammatory proteins, including cyclooxygenase-2 (Cox-2), Nox2, and vascular cell adhesion molecule-1 (VCAM-1), and infiltration of Nox2-containing T lymphocytes into the infarcted brain, whereas such changes are minimal in female mice after ischemia–reperfusion (I-R). Infarct volume after I-R was no greater at 72 h than at 24 h in either gender. Infarct development was Nox2 dependent in male but not in female mice, and Nox2 within the infarct was predominantly localized in T lymphocytes. Stroke resulted in an ∼15-fold increase in Nox2-dependent superoxide production by circulating, but not spleen-derived, T lymphocytes in male mice, and this was ∼sevenfold greater than in female mice. These circulating immune cells may thus represent a major and previously unrecognized source of superoxide in the acutely ischemic and reperfused brain of males (and potentially in postmenopausal females). Our findings provide novel insights into mechanisms that could be therapeutically targeted in acute ischemic stroke patients who receive thrombolysis therapy to induce cerebral reperfusion.

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Infarct Volume Predicts Delayed Recovery in Patients with Subarachnoid Hemorrhage and Severe Neurological Deficits

Neurocritical Care ◽

10.1007/s12028-013-9869-3 ◽

2013 ◽

Vol 19 (3) ◽

pp. 293-298 ◽

Cited By ~ 8

Author(s):

Neil F. Rosenberg ◽

Storm M. Liebling ◽

Adam R. Kosteva ◽

Matthew B. Maas ◽

Shyam Prabhakaran ◽

...

Keyword(s):

Subarachnoid Hemorrhage ◽

Infarct Volume ◽

Neurological Deficits ◽

Delayed Recovery

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Oral prescription opioid-seeking behavior in male and female mice

10.1101/621177 ◽

2019 ◽

Author(s):

Alysabeth G. Phillips ◽

Dillon J. McGovern ◽

Soo Lee ◽

Kyu Ro ◽

David T. Huynh ◽

...

Keyword(s):

Sex Differences ◽

Oral Drug ◽

Prescription Opioid ◽

Male Mice ◽

Self Administration ◽

Female Mice ◽

Seeking Behavior ◽

Opioid Users ◽

Psychomotor Stimulation

AbstractA significant portion of prescription opioid users self-administer orally rather than intravenously. Animal models of opioid addiction have demonstrated that intravenous cues are sufficient to cause drug-seeking. However, intravenous models may not model oral users, and the preference to self-administer orally appears to be partially influenced by the user’s sex. Our objectives were to determine whether oral opioid-associated cues are sufficient for relapse and whether sex differences exist in relapse susceptibility. Mice orally self-administered escalating doses of oxycodone under postprandial (prefed) or non-postprandial (no prefeeding) conditions. Both sexes demonstrated robust cue-induced reinstatement. In separate mice we found that oral oxycodone cues were sufficient to reinstate extinguished oral oxycodone-seeking behavior in the absence of postprandial or prior water self-administration training. During self-administration, we found that female mice earned significantly more mg/kg oxycodone than male mice. Follow-up studies indicated sex differences in psychomotor stimulation and plasma oxycodone/oxymorphone following oral oxycodone administration. In addition, gonadal steroid studies were performed in which we found divergent responses where ovariectomy enhanced and orchiectomy suppressed oral self-administration. While the suppressive effects of orchiectomy were identified across doses and postprandial conditions, the enhancing effects of ovariectomy were selective to non-postprandial conditions. These studies establish that 1) oral drug cues are sufficient to cause reinstatement that is independent of prandial conditions and water-seeking behavior, 2) earned oral oxycodone is larger in female mice compared with male mice potentially through differences in psychomotor stimulation and drug metabolism, and 3) gonadectomy produces divergent effects on oral oxycodone self-administration between sexes.

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Sex differences in nociceptor translatomes contribute to divergent prostaglandin signaling in male and female mice

10.1101/2020.07.31.231753 ◽

2020 ◽

Author(s):

Diana Tavares-Ferreira ◽

Pradipta R. Ray ◽

Ishwarya Sankaranarayanan ◽

Galo L. Mejia ◽

Andi Wangzhou ◽

...

Keyword(s):

Sex Differences ◽

Cortical Neurons ◽

Affinity Purification ◽

Male Mice ◽

Protective Behaviors ◽

Pain Mechanisms ◽

Female Mice ◽

Mechanistic Basis ◽

High Doses ◽

Neuronal Functions

ABSTRACTBackgroundThere are clinically relevant sex differences in acute and chronic pain mechanisms, but we are only beginning to understand their mechanistic basis. Transcriptome analyses of rodent whole dorsal root ganglion (DRG) have revealed sex differences, mostly in immune cells. We examined the transcriptome and translatome of the mouse DRG with the goal of identifying sex differences.MethodsWe used Translating Ribosome Affinity Purification (TRAP) sequencing and behavioral pharmacology to test the hypothesis that nociceptor (Nav1.8 expressing neurons) translatomes would differ by sex.ResultsWe found 66 genes whose mRNA were sex-differentially bound to nociceptor ribosomes. Many of these genes have known neuronal functions but have not been explored in sex differences in pain. We focused on Ptgds, which was increased in female mice. The mRNA encodes the prostaglandin D2 (PGD2) synthesizing enzyme. We observed increased Ptgds protein and PGD2 in female mouse DRG. The Ptgds inhibitor AT-56 caused intense pain behaviors in male mice but was only effective at high doses in females. Conversely, female mice responded more robustly to another major prostaglandin, PGE2, than did male mice. Ptgds protein expression was also higher in female cortical neurons, suggesting DRG findings may be generalizable to other nervous system structures.ConclusionsNociceptor TRAP sequencing (TRAP-seq) reveals unexpected sex differences in one of the oldest known nociceptive signaling molecule families, the prostaglandins. Our results demonstrate that translatome analysis reveals physiologically relevant sex differences important for fundamental protective behaviors driven by nociceptors.

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Abstract 113: Regulatory T Cells Contribute to Sexual Dimorphic Outcomes After Acute Ischemic Brain Injury

Stroke ◽

10.1161/str.51.suppl_1.113 ◽

2020 ◽

Vol 51 (Suppl_1) ◽

Author(s):

Zeyu Sun ◽

Wei Su ◽

Ligen Shi ◽

Jie Chen ◽

Xiaoming Hu

Keyword(s):

T Cells ◽

Regulatory T Cells ◽

Immune Cell ◽

Infarct Volume ◽

Young Male ◽

Young Female ◽

Male Mice ◽

Stroke Outcomes ◽

Female Mice ◽

Ifn Γ

Introduction: The contribution of CD4 + Foxp3 + regulatory T cells (Treg) to acute stroke outcomes has been controversially reported in young male murine models of stroke, their effects in female stroke mice, however, are not characterized. This study explored the sexual dimorphisms in Treg and its contribution to acute stroke outcomes. Methods: Cerebral ischemia was induced by 60 min tMCAO in male or female (Young: 10-week, aged: 15-month) wild type (WT) mice and DTR mice expressing the diphtheria toxin (DT) receptor under the control of Foxp3 promoter. Tregs depletion was achieved by DT injection in DTR mice for 3 days prior to tMCAO. Infarct volume, sensorimotor functions and peripheral immune cell populations were assessed up to 5d after stroke. For RNA sequencing analysis, Tregs were sorted from blood of male or female DTR mice at 5d after sham or tMCAO surgery. Results: Young Treg competent (WT mice or DTR mice without DT) female mice exhibited significantly reduced infarct volume, as assessed by MRI T2 scanning and MAP2 staining, and greatly improved sensorimotor functions (rotarod test and adhesive removal test) compared to age- and genotype-matched male mice (n=8/group) 5d after tMCAO. Treg depletion deprived the neuroprotection in young female, while showed no significant effect on young male or aged female mice (n=8/group). RNA-seq analysis showed that IFN-γ signaling was downregulated in female Treg while upregulated in male Treg, suggesting a sexual difference in Treg-mediated immune response after stroke. Flow cytometry revealed ameliorated immune cell activation in blood and brain in female vs male mice 5d after stroke. Furthermore, Treg were isolated from young female, young male, aged female or female mice subjected to ovariectomy, and adoptively transferred (1 million cell/animal, iv) to young male mice 1 hour after tMCAO. Only young female Treg significantly reduced the infarct volume and improved sensorimotor functions compared to other treatment groups (n=7-8/group). Conclusion: Treg contribute to the neuroprotection in young female vs male in an age- and hormone-dependent manner. Transcriptomic analysis uncovered sexual differences in an IFN-γ centered regulatory pathways in Tregs, which keep post-stroke immune responses in check.

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Abstract WP308: Role of Microthrombi and Vasospasm in the Development of Delayed Neurological Deficits After Subarachnoid Hemorrhage in Mice

Stroke ◽

10.1161/str.51.suppl_1.wp308 ◽

2020 ◽

Vol 51 (Suppl_1) ◽

Author(s):

Devin W McBride ◽

Ari Dienel ◽

Remya A Veettil ◽

Kanako Matsumura ◽

Peeyush Kumar T. ◽

...

Keyword(s):

Subarachnoid Hemorrhage ◽

Mouse Model ◽

Experimental Studies ◽

Brain Slices ◽

Neurological Deficits ◽

Male Mice ◽

Male And Female ◽

Delayed Neurological Deterioration ◽

First Time

Rationale: Microthrombosis has been suggested as a major factor contributing to delayed neurological deterioration in patients after subarachnoid hemorrhage (SAH). However, experimental studies on the role of microthrombi in delayed deficits after SAH has not been investigated. Our hypothesis is that, following SAH, mice which develop delayed neurological deficits have a greater number of microthrombi than mice which do not develop delayed neurological deficits. Methods: SAH was induced in adult male and female C57BL/6 mice via endovascular perforation. Mice were randomly assigned into sham (n=6/sex) or SAH groups (n=22-24/sex). Neurobehavior was performed on days 1-3, 5, and 7 post-SAH using a composite neuroscore. Animals were sacrificed on the day of delayed deficits or 7 days post-SAH. Microthrombi count and vessel diameters (for vasospasm) were measured using H&E stained brain slices. All outcomes were performed and all data were analyzed by a blinded investigator. Results: Seventeen percent (4/24) of male mice and thirty-six percent (8/22) of female mice developed delayed deficits on days 3-5 post-SAH (Figures 1A and 1B). Those mice which developed delayed deficits had significantly more microthrombi in their brains than mice which did not develop delayed deficits; vasospasm did not correlate with delayed deficits. Additionally, female SAH mice develop delayed deficits at a higher frequency than males (Figure 1C). Conclusions: This work shows for the first time delayed deficits in a SAH mouse model. Further, microthrombi correlated with delayed deficits, whereas no correlation was between delayed deficits and vasospasm. The data within this study suggests that preventing microthrombi may improve functional recovery and reduce the risk of delayed deficits.

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Sex differences in adaptive downregulation of pre-macula densa sodium transporters with ANG II infusion in mice

AJP Renal Physiology ◽

10.1152/ajprenal.00088.2009 ◽

2010 ◽

Vol 298 (1) ◽

pp. F187-F195 ◽

Cited By ~ 12

Author(s):

Swasti Tiwari ◽

Lijun Li ◽

Shahla Riazi ◽

Veerendra K. Madala Halagappa ◽

Carolyn M. Ecelbarger

Keyword(s):

Sex Differences ◽

Plasma Testosterone ◽

Sodium Reabsorption ◽

Thick Ascending Limb ◽

Ang Ii ◽

Male Mice ◽

Male And Female ◽

Female Mice ◽

Sodium Transporters ◽

Pressure Natriuresis

An increase in blood pressure (BP) due to angiotensin II (ANG II) infusion or other means is associated with adaptive pressure natriuresis due to reduced sodium reabsorption primarily in proximal tubule (PT) and thick ascending limb (TAL). We tested the hypothesis that male and female mice would show differential response to ANG II infusion with regard to the regulation of the protein abundance of sodium transporters in the PT and TAL and that these responses would be modulated by aging. Young (∼3 mo) and old (∼21 mo) male and female mice were infused with ANG II at 800 ng·kg body wt−1·min−1 by osmotic minipump for 7 days or received a sham operation. ANG II increased mean arterial pressure (MAP), measured by radiotelemetry, significantly more in male mice of both ages (increased ∼30–40 mmHg), compared with females (increased ∼15–25 mmHg). On day 1, MAP was also significantly increased in old mice, relative to young ( P = 0.01). ANG II infusion was associated with a significant decline in plasma testosterone (to <30% of control male) in male mice and rise in young female mice (to 478% of control female). No sex differences were found in the upregulation of the sodium hydrogen exchanger abundance on Western blots observed with ANG II infusion or the downregulation of the sodium phosphate cotransporter; however, aging did impact on some of these changes. Male mice (especially young) also had significantly reduced levels of the TAL bumetanide-sensitive Na-K-2Cl cotransporter (to 60% of male control), while young females showed an increase (to 126% of female control) with ANG II infusion. These sex differences do not support impaired pressure natriuresis in male mice, but might reflect a greater need and attempt to mount an appropriately BP-metered natriuretic response by additional downregulation of TAL sodium reabsorption.

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