Gender affects macrophage cytokine and prostaglandin E2 production and PGE2 receptor expression after trauma1

doi:10.1016/j.jss.2004.04.020

Journal of Surgical Research

Volume 122, Issue 1, November 2004, Pages 1-7

https://doi.org/10.1016/j.jss.2004.04.020 Get rights and content

Background

Gender influences morbidity and mortality after injury. Hormonal differences are important; however, the role of prostaglandins as mediators in immune dysfunction relating to gender differences after trauma is unclear. We hypothesized that gender-dependent differences in PGE₂ receptor expression and signaling may be involved in immune-related differences. This study determined prostaglandin receptor subtype (EP1–EP4) expression following injury and determined whether gender differences influence EP receptor expression.

Materials and methods

BALB/c male and female mice (estrus and pro-estrus) (n = 6 per group) were subjected to femur fracture and 40% hemorrhage (trauma) or sham injury (anesthesia). Seven days later, the splenic macrophages were harvested and stimulated with lipopolysaccharide (Escherichia coli serotype O55:B5). After 6 h mRNA samples were collected for EP receptor mRNA expression and at 24 h supernatants were collected for PGE₂, TNF-α, and IL-6 production.

Results

The expression of EP2-4 receptors was higher in female pro-estrus mice compared with male mice. EP1 receptor expression was higher in males than pro-estrus females. There was decreased expression of all four receptors after trauma in female estrus compared with control estrus mice. Macrophage PGE₂, TNF-α, and IL-6 production was significantly increased in injured female mice compared with female controls but there were no differences in injured male mice compared with male controls. PGE₂ and TNF-α production by traumatized male mice were significantly less than that produced by traumatized pro-estrus females.

Conclusions

These data suggest gender-related differences in response to traumatic injury and that alterations in specific EP receptor subtypes may be involved in immune dysfunction after injury. Studies to evaluate targeted modulation of these receptor subtypes may provide further insights to gender-specific differences in the immune response after injury.

Introduction

Major injury leads to impaired immune responses and increases the risk of subsequent infectious complications and it has been suggested that gender-related differences may affect outcome following trauma [1, 2]. Several studies purport that severely injured males have an increased propensity to immune suppression and morbidity compared with similarly injured females [3, 4]. Knoferl et al. [5] suggested that female sex hormones may be beneficial in preventing trauma-induced immune depression and increased susceptibility to subsequent sepsis, while Angele et al. reported androgen receptor blockade to be beneficial after major injury in mice [6].

The immunosuppressive events that predispose the injured patient to this susceptibility have been associated with a persistence of prostaglandins, proinflammatory cytokines, and other mediators that lead to end-organ damage and/or infections that ultimately may cause multiple organ failure and death [7]. In particular, prostaglandin E₂ (PGE₂) produced by stimulated macrophages is significantly elevated in many immune compromised states in humans and animal models, including burns, sepsis, and shock [8, 9, 10, 11, 12]. It is an early mediator released following injury, making it a target for modulation of the immune response [13].

There is a body of literature with regard to sexual dimorphism and immune function following major injury and onset of infection. Numerous groups have shown that in ICU settings women with sepsis have a survival advantage over men [14, 15]. Experimental models suggest that the pro-estrus state affords protection to traumatized female mice compared with males having suffered an identical insult [3, 16]. Yet despite strong evidence to suggest outcome advantage mediated by female sex hormones, other studies show that injured females have elevated circulating levels of PGE₂ [17, 18] and the immune suppressive effects of PGE₂ are well documented [9, 19, 20, 21]. The results from these studies, when considered with previous findings in our murine model of trauma and hemorrhage as well as those from other laboratories using various models of injury, suggest detrimental effects of PGE₂ [22, 23, 24].

With regard to female mice, the differences in morbidity following trauma are related to the hormonal phase [25]; however, no data exist in relation to gender-related PGE₂ receptor subtypes after trauma. Recent characterization of specific subtypes of the prostaglandin receptors (EP1–EP4) has aided understanding of the complexity of PGE₂ signaling. All four receptor subtypes are activated by PGE₂, but each functions via a different intracellular signaling pathway. EP2 and EP4 receptors are preferentially present in the thymus, lung, and spleen and may be more involved in immune cell activation than EP1 and EP3 receptors [26]. This led us to speculate that certain EP receptor subtypes may play a differential role in immune regulation.

No studies have yet examined the relationship between gender and EP receptor subtype expression after trauma. We therefore aimed to evaluate the following: (1) the presence and distribution of specific EP receptors in splenic macrophages from male and female mice, (2) the effect of trauma on EP receptor expression in mice, and (3) if gender (male versus female) or hormonal status (pro-estrus versus estrus female) causes different patterns of EP receptor expression after trauma. With these aims in mind, we propose to define the patterns of PGE₂ receptor subtypes and suggest their importance in explaining altered immune responses after injury in both males and females.

Section snippets

Animals

Inbred, young (6–8 weeks old) female BALB/c mice (weight, 19–22 g) were purchased from Charles River Laboratories (Wilmington, MA). On arrival at the housing facility, animals were acclimatized for 2–4 days prior to experiments and allowed standard laboratory chow and water ad libitum. The Association for Assessment and Accreditation of Laboratory Animal Care approved this facility and all protocols were approved by the Institutional Animal Care and Use Committee of Weill Medical College of

Splenic macrophage eicosanoid and cytokine production

Figure 1 shows PGE₂ production by male and pro-estrus female mice at 7 days after trauma. There was no difference in splenic macrophage PGE₂ production in response to trauma in male mice compared with control males (61.81 ± 5.85 versus 48.08 ± 6.83 pg/μg protein, TM versus CM). Splenic macrophages from traumatized pro-estrus female mice however produced significantly greater levels of PGE₂ compared with their respective controls (103.02 ± 7.8 versus 33.73 ± 7.2 pg/μg protein, TF versus CF, P <

Discussion

This study demonstrates gender-specific differences in the response to traumatic injury. We have shown that splenic macrophage function is muted in male mice compared with females after trauma. The combined injury of femur fracture and hemorrhage only moderately affected splenic macrophage function in male mice, although significantly elevated PGE₂, TNF-α, and IL-6 levels were observed from splenic macrophages isolated from female mice subjected to an identical insult. Within the female group

Uncited references

This section comprises references that occur in the reference list but not in the body of the text. Please position each reference in the text or delete it. Any references not dealt with will be retained in this section: [34].

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Cited by (19)

Sex-biased eicosanoid biology: Impact for sex differences in inflammation and consequences for pharmacotherapy
2017, Biochemical Pharmacology
Citation Excerpt :
Deletion of the PGI2 receptor abolished atheroprotective effects of estrogen in ovariectomized female mice [55]. Following traumatic injury, the expression of PGE2 receptors EP2-4 was higher in female pro-estrus versus male mice, EP1 receptor expression was higher in males, and all four PGE2 receptors were decreased after trauma in female estrus compared with control estrus mice [56]. Sex differences in prostanoid biosynthesis and modulation by sex hormones were also studied at the cellular level in vitro.
The incidence, severity and progression of autoimmune diseases (e.g. scleroderma, multiple sclerosis, rheumatoid arthritis) and certain inflammatory diseases (e.g. asthma) are sex-biased where these pathologies dominate in women. However, other immune disorders such as sepsis, post-surgery infections and gout display higher incidence and severity in men. The molecular and cellular basis underlying this sex dimorphism remains incompletely elucidated but may provide important insights for sex-specific pharmacotherapy. Nevertheless, the sex as a variable in biochemical and preclinical research on inflammation is often neglected. Thus, respective animal studies are routinely performed with males, and experiments with isolated cells rarely report the sex of the donor. However, sex differences on the cellular level do exist, in particular related to inflammatory processes that prompt for sex-specific appreciation of inflammation research. For instance, the biosynthesis of pro-inflammatory eicosanoids is sex-biased where leukotriene (LT) formation is under control of testosterone that regulates the subcellular localization of the key enzyme 5-lipoxygenase, with possible implications for gender-tailored pharmacotherapy of LT-related disorders (i.e. asthma). Moreover, prostaglandin (PG) production is sex-biased, and sex-dependent efficacy of aspirin was evident in several clinical trials. Here, we highlight the sex bias in eicosanoid biology possibly underlying the obvious sex disparities in inflammation, stimulating scientists to take sex into account when studying the pathophysiology and pharmacotherapy of inflammatory diseases.
Automobile versus pedestrian injuries: Does gender matter?
2011, Journal of Emergency Medicine
Citation Excerpt :
Women are generally shorter than men and, as a result, the pelvis is more likely to suffer a direct hit during AVP trauma. On the other hand, in taller men, the force of bumper impact is more likely to be discharged on the femur or tibia (2,8). In a previous study from our center, the incidence of pelvic fractures was twice as common in adults compared to pediatric patients after AVP trauma (15.8% vs. 6.7%, p < 0.0001) (17).
Background: Automobile vs. pedestrian (AVP) injuries cause substantial morbidity and mortality. Gender may be an important factor in determining the anatomic distribution and severity of these injuries. The objective of this study was to examine the effect of gender on the nature and severity of automobile vs. pedestrian injuries and the outcome. Methods: Trauma registry study that included all AVP pedestrian injuries admitted during a 14-year period to a Level I trauma center. The following variables were included in an Excel (Microsoft Corporation, Redmond, WA) file for the purpose of this study: age, gender, body area Abbreviated Injury Score, Injury Severity Score, specific fractures (pelvic, spine, femur, tibia), survival, and intensive care unit (ICU) and hospital length of stay. Results: The study population included 6965 patients, 67.3% of whom were male. Overall, 20.7% were in the age group < 15 years, 60.5% in the age group 15–55 years, 7.6% in the age group 56–65 years, and 11.1% in the age group > 65 years. Pelvic fractures were significantly more common in females than males (20.7% vs. 11.4%, respectively, p < 0.0001). This difference was present in all age groups, but especially in the groups 56–65 years (28.5% vs. 12.3%, respectively, p < 0.0001) and > 65 years (32.5% vs. 15.7%, respectively, p < 0.0001). Males in the age group 15–55 years were significantly more likely to suffer tibia fractures (31.8% vs. 25.7%, respectively, p < 0.001). Multivariate analysis showed no difference in survival or ICU stay between the two genders, but there was a significantly longer hospital stay in males 15–65 years. Conclusions: Gender plays a significant role in the incidence of pelvic and tibial fractures but has no effect on survival or ICU stay, but male patients in the age group 15–65 years had a significantly longer hospital stay.
Association study of EP1 gene polymorphisms with suicide completers in the Japanese population
2011, Progress in Neuro-Psychopharmacology and Biological Psychiatry
Citation Excerpt :
Animal studies revealed that PGE2 signaling is involved in brain gender differentiation and subsequent disruption might cause behavioral abnormalities later in life (Ottem et al., 2004). EP1 expression is also related to hormonal status in females, and EP1 signaling might induce different actions in males and females (Stock et al., 2000; Stapleton et al., 2004). In our study, the MAFs of the female suicide completers were lower than the female controls in all analyses resulting in nominal significant difference.
Both environmental and genetic factors have been reported to be involved in suicidal behaviors. Considerable evidence indicates that impulsive aggression is one of the important risk factors that contribute to suicide. A recent study has shown that prostaglandin E2 type 1 receptor (EP1) signaling regulates impulsive-aggressive behaviors in mice under both social and environmental stresses. To test the possible involvement of the EP1 gene in suicide, we carried out an association study of EP1 gene polymorphisms with suicide completers in the Japanese population.
We studied 5 SNPs including one SNP in exon 2 (rs3745459) and four SNPs in the potential promoter region of the EP1 gene (rs3810255, rs3810254, rs3810253 and rs10416814) in 374 healthy control and 287 completed suicide victims using standard Taqman probe genotyping assays.
No significant differences of the genotypic distribution, allelic frequency or haplotype distribution between controls and suicide completers were found. Gender based analysis revealed that genotypic, allelic and haplotypic distributions of rs3810255, rs3810254, rs3810253 and rs10416814 SNPs were significantly different between the female control and female suicide groups, although the differences did not withstand correction for multiple comparisons.
We could not find an association of EP1 gene with suicide in the Japanese population. Because several SNPs in the promoter region of the EP1 gene were nominally significantly associated with suicide in the female, further studies with a larger sample size and different population are needed to confirm this result.
β<inf>2</inf> Adrenoreceptor blockade attenuates the hyperinflammatory response induced by traumatic injury
2009, Surgery
Major operative injury initiates immunologic changes that may result in a systemic inflammatory response, leading to organ dysfunction/sepsis. Catecholamines seem to be key mediators. The effects of β₂ receptor blockade in vitro and in vivo before and after operative injury were studied to clarify their role.
In vitro studies were done in RAW 264.7 cells using epinephrine (50 μmol/L) with or without α₂- and β₂-receptor blockade. Comparative gene expression analysis on the Toll-like receptor (TLR)-4 receptor signaling pathway was performed between RAW cells pretreated with epinephrine with subsequent lipopolysaccharide (LPS) stimulation versus LPS alone. Confirmatory studies were performed by real-time reverse transcription polymerase chain reaction (RT-PCR). Tumor necrosis factor (TNF)-α gene and protein expression were determined via real time RT-PCR and enzyme-linked immunosorbent assay, respectively. In vivo, Balb/C mice received no treatment nor injury (group I). Group II received operative injury and vehicle injection, group III received ICI 118,551 (β₂-receptor antagonist) 30 minutes before or in separate studies after operative injury. At 7 days, splenic macrophages were harvested and cytokine production was measured with or without LPS. In separate experiments, cecal ligation and puncture (CLP) was done 7 days after operation and survival determined.
In vitro studies demonstrated that epinephrine pretreatment significantly increased TNF-α production with LPS stimulation (P < .05). β₂-Receptor blockade significantly attenuated (P < .05) LPS stimulated TNF-α production. MD-2, an essential coactivator of TLR-4 signaling, gene expression was significantly elevated when cells were pretreated with epinephrine before LPS exposure (P < .001). In vivo studies demonstrated a significant decrease (P < .05) in TNF-α and interleukin-6 production in the ICI 118,551 group. Similar findings were demonstrated measuring monocyte chemoattractant protein-1 and interferon-γ cytokine levels (P < .05) versus no treatment. ICI 118,551 treatment 30 minutes before operation demonstrated a 31% reduction in mortality after CLP (P < .05).
This study demonstrates that β₂-receptor blockade reduces macrophage cytokine production and improves survival showing the critical importance of catecholamines to the immunologic response in surgery.
Sex differences and estrogen modulation of the cellular immune response after injury
2008, Cellular Immunology
Citation Excerpt :
Other trauma–hemorrhage studies show that macrophage production of PGE2, TNF-α, and IL-6 was significantly increased in injured female mice compared with female controls, but there were no differences in injured male mice compared with sex-matched controls. PGE2 and TNF-α production by male mice following traumatic injury was significantly less than that produced by injured proestrus females [71]. Due to the dichotomy in the literature, further studies need to be performed to further understand the role of PGE2 in the immune response following trauma.
Cell-mediated immunity is extremely important for resolution of infection and for proper healing from injury. However, the cellular immune response is dysregulated following injuries such as burn and hemorrhage. Sex hormones are known to regulate immunity, and a well-documented dichotomy exists in the immune response to injury between the sexes. This disparity is caused by differences in immune cell activation, infiltration, and cytokine production during and after injury. Estrogen and testosterone can positively or negatively regulate the cellular immune response either by aiding in resolution or by compounding the morbidity and mortality. It is apparent that the hormonal dysregulation is dependent not only on the type of injury sustained but also the amount of circulating hormones. Therefore, it may be possible to design sex-specific therapies to improve immunological function and patient outcome.
Association between prostaglandin E2 receptor gene and essential hypertension
2007, Prostaglandins Leukotrienes and Essential Fatty Acids
Essential hypertension (EH) is a complex multifactorial polygenic disorder that is thought to result from an interaction between an individual's genetic makeup and various environmental factors. In the kidney, prostaglandins (PGs) are important mediators of vascular tone and salt and water homeostasis, and are involved in the mediation and/or modulation of hormonal action. In previous studies, mice deficient in the prostaglandin E2 (PGE₂) EP2 receptor had resting systolic blood pressure (BP) that was significantly lower than that of wild-type controls. The BP of those mice increased when they were put on a high-salt diet, suggesting that the EP2 receptor is involved in sodium handling by the kidney. In the present study, we investigated the association between EH and nucleotide polymorphisms in the gene encoding the prostaglandin E2 receptor subtype EP2 (PTGER2).
We selected three single-nucleotide polymorphisms (SNP) in the human PTGER2 gene (rs1254601, rs2075797, and rs17197), and we performed a genetic association study of 266 EH patients and 253 age-matched normotensive (NT) controls.
There was no significant difference in overall distribution of genotypes or alleles of any of the SNP between the EH and NT groups. However, among men, the A/A type of the SNP rs17197 (rs17197, A/G in 3′UTR) was significantly more frequent in EH subjects than in NT subjects (P=0.041).
The present findings suggest that rs17197 is useful as a genetic marker of EH in men.

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¹: This study was supported by NIH Grant 1R01 AI48755-01A2 (J.M.D.) and NIH Training Grant T32 GM08466-06 (J.M.D.).

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Shock/Sepsis/Trauma/Critical CareGender affects macrophage cytokine and prostaglandin E2 production and PGE2 receptor expression after trauma11

Background

Materials and methods

Results

Conclusions

Introduction

Section snippets

Animals

Splenic macrophage eicosanoid and cytokine production

Discussion

Uncited references

Cytokine

Surgery

Surgery

Immunol. Lett.

Surgery

Cell Immunol.

J. Surg. Res.

Cytokine

Male gender is a risk factor for major infections after surgery

Arch. Surg.

Sex steroids regulate pro- and anti-inflammatory cytokine release by macrophages after trauma-hemorrhage

Am. J. Physiol.

Females in proestrus state maintain splenic immune functions and tolerate sepsis better than males

Crit. Care Med.

Female sex hormones regulate macrophage function after trauma-hemorrhage and prevent increased death rate from subsequent sepsis

Ann. Surg.

Testosterone receptor blockade after hemorrhage in males. Restoration of the depressed immune functions and improved survival following subsequent sepsis

Arch. Surg.

The role of prostaglandin E2 in immune suppression following injury

Ann. Surg.

Prostaglandin E2 (PGE2)-dependent suppression of interleukin-2 production in patients with major trauma

J. Trauma

Chemically induced hypotension increases PGE2 release and depresses macrophage antigen presentation

Am. J. Physiol.

Time course for production of cytokines and prostaglandin E2 by macrophages isolated after thermal injury and bacterial translocation

Circ. Shock

Incidence of septic complications and multiple organ failure in severely injured patients is sex specific

J. Trauma

Gender differences in human sepsis

Arch. Surg.

Improved survival in uncontrolled hemorrhagic shock induced by massive splenic injury in the proestrus phase of the reproductive cycle in the female rat

Shock

Update on the mechanisms of immune suppression of injury and immune modulation

World J. Surg.

Shock/Sepsis/Trauma/Critical Care
Gender affects macrophage cytokine and prostaglandin E₂ production and PGE₂ receptor expression after trauma¹1

Prostaglandin E2 (PGE₂)-dependent suppression of interleukin-2 production in patients with major trauma

Chemically induced hypotension increases PGE₂ release and depresses macrophage antigen presentation