The Experimental Research Of Gender Dimorphism In Poly-Morphonuclear Cell Response And Metoclopramide Intervention Following Trauma-Hemorrhagic Shock

ObjectiveAlthough gender influences T-cell, macrophage and organ functions following trauma-hemorrhage and resuscitation, it remains unknown whether it also influences polymorphonuclear cell (PMN) activity under such conditions. The aim of this series of study including:1. To establish a non-heparinized model of trauma-hemorrhage in the rat, and determine the gender dimorphism of circulating PMN's response to trauma-hemorrhage and resuscitation.2. To determine whether the behavior of polymorphonuclear leukocyte adhesion to the endothelial cell after trauma-hemorrhage and resuscitation is different between genders by means of intravital microscopy. The expression of cell adhesion molecule CD11b of PMN was determined too.3. To determine whether administration of metoclopramide (MCP) -a drug can increases the anterior pituitary secrete prolactin produces any beneficial effects on the depressed immune function and the delayed apoptosis of PMN after severe hemorrhage.Methods1. Proestrus female and adult male Sprague-Dawley rats (275 - 325g) were divided into 3 groups: Female, Male and Contrast. After anaesthesia a 5-cmmidline laparotomy was made to make soft tissue injury and then the wound were closed in layers. After that the rats were bled to a mean arterial pressure (MAP) of 35±5mmHg for lh, and then the animals were resuscitated with 4 times the volume of withdrawn blood with Ringer's lactate over 60 min. The contrast group underwent no shock and no treatment. Additionally, total shed blood was reinfused. 2h and 24h after resuscitation circulating PMN were assessed for superoxide (O2") and elastase production and tissues were analyzed for myeloperoxidase (MPO) activity and MDA (Malondialdehyde) as a marker of oxidative injury.2. 2h and 24h after hemorrhagic shock resuscitation a 5-cm midline laparotomy was made and the mesentery near to the ileum was exteriorized and the microcirculation blood flow as well as PMN-endothelial cell interaction was evaluated by means of intravital microscopy. The number of leukocytes sticking to the venular endothelium was determined and the expression of cell adhesion molecule CD1 lb of PMN was determined by flow cytometry.3. Male Sprague-Dawley rats (275-325g) were made 5-cm midline laparotomy and bled to a mean arterial pressure of 35±5mmHg for lh, then adequately resuscitated and randomized into 3 groups: One group received metoclopramide (lOOmg/lOOg body weight, subcutaneously) before resuscitation;the second group were treated with same volume of saline vehicle;the contrast group underwent no shock and no treatment. 2h after saline or MCP injection, the animals were killed and spleen tissue was homogenated used to determine IL-lp and IL-6. Plasma TNF-a and IL-8 levels were also measured. The expression of PMN Fas was measured too by flow cytometry.Results1. At 2 and 24 h after resuscitation, PMA stimulated O2" production was notinfluenced by T-H or gender. In contrast, fMLP-stimulated O2" and LPS-stimulated elastase release by PMNs from male T-H rats were greater than that of female rats. A significant MPO activity and MDA in tissues of both male and female rats were induced;however, MPO activity and MDA levels were higher in males following T-H.2. 2h after treatment, Female (1.8±0.3), and Contrast (0.4 ± 0.2) groups presented significant reductions in leukocyte adherence (cells/1 OOjum venule length), compared with Male group (4.2±0.9, P< 0.05). No differences were observed 6h after treatment on leukocyte adherence. The expression of CD1 lb on PMN was significantly higher in Male animals compared with the Female and Contrast groups (P< 0.01).3. Release of IL-ip and IL-6 by spleen tissue from metoclopramide treated group was maintained after trauma-hemorrhage, whereas release of IL-ip and IL-6 by spleen tissue from saline treated group rat was depressed by approximately 55%. In contrast, trauma-hemorrhage resulted in a threefold increase of plasma lever of TNF-a and IL-8 in saline treated group and plasma concentrations of TNF-a and IL-8 were unchanged in metoclopramide treated rat under such conditions. The Fas expression of PMN was depressed in saline treated group (P<0.01) than the Contrast group, although the Fas expression of PMN was also depressed in metoclopramide treated rats (P<0.05), but it was also higher than that of saline treated group.Conclusion1. Decreased PMN priming and activation in proestrus females, compared to males, occurs following T-H resulting in decreased cellular injury and organ damage that is likely to contribute to improved outcome under those conditions.2. The PMN-endothelial cell interactions and PMN CD lib expression were down regulated in female rats after hemorrhagic shock and resuscitation.3. The immunosuppression after hemorrhage may be restored by the use of single dose metoclopramide. Furthermore, metoclopramide may be a useful adjuvant in the treatment of the trauma-hemorrhagic shock-induced immunosuppression.