Impact of 5-lipoxygenase inhibitors on the spatiotemporal distribution of inflammatory cells and neuronal COX-2 expression following experimental traumatic brain injury in rats.

The inflammatory response following traumatic brain injury (TBI) contributes to neuronal death with poor outcome. Although anti-inflammatory strategies were beneficial in the experimental TBI, clinical translations mostly failed, probably caused by the complexity of involved cells and mediators. We recently showed in a rat model of controlled cortical impact (CCI) that leukotriene inhibitors (LIs) attenuate contusion growth and improve neuronal survival. This study focuses on spatiotemporal characteristics of macrophages and granulocytes, typically involved in inflammatory processes, and neuronal COX-2 expression. Effects of treatment with LIs (Boscari/MK-886), started prior trauma, were evaluated by quantifying CD68(+), CD43(+) and COX-2(+) cells 24h and 72 h post-CCI in the parietal cortex (PC), CA3 region, dentate gyrus (DG) and visual/auditory cortex (v/aC). Correlations were applied to identify intercellular relationships. At 24h, untreated animals showed granulocyte invasion in all regions, decreasing towards 72 h. Macrophages increased from 24h to 72 h post-CCI in PC and v/aC. COX-2(+) neurones showed no temporal changes, except of an increase in the CA3 region at 72 h. Treatment reduced granulocytes at 24h in the pericontusional zone and hippocampus, and macrophages at 72 h in the PC and v/aC. COX-2 expression remained unaffected by LIs, except of time-specific changes in the DG (increase/decrease at 24/72 h). Interrelations confirmed concomitant cellular reactions beyond the initial trauma site. In conclusion, LIs attenuated the cellular inflammatory response following CCI. Future studies have to clarify region-specific effects and explore the potential of a clinically more relevant therapeutic approach applying LIs after CCI.

Early inflammatory response of knee ligaments to prolotherapy in a rat model.

Prolotherapy is an alternative injection-based therapy for chronic musculoskeletal pain. Three different proliferants, D-glucose (dextrose), phenol-glucose-glycerine (P2G), and sodium morrhuate, used in prolotherapy are hypothesized to strengthen and reorganize chronically injured soft tissue and decrease pain through modulation of the inflammatory process. Our hypothesis is that commonly used prolotherapy solutions will induce inflammation (leukocyte and macrophage infiltration) in medial collateral ligaments (MCLs) compared to needlestick, saline injection, and no-injection controls. MCLs of 84 Sprague- Dawley rats were injected one time at both the tibial and femoral insertions. Immunohistochemistry (IHC) was used to determine the inflammatory response at three locations (tibial and femoral insertions and midsubstance) 6, 24, and 72 h after dextrose injection compared to saline- and no-injection controls and collagenase (positive control) (n = 4). qPCR was used to analyze gene expression 24 h postinjection (n = 4). Sodium morrhuate, P2G, and needlestick control were also investigated after 24 h (n = 4). In general, inflammation (CD43+, ED1+, and ED2+ cells) increased after prolotherapy injection compared to no-injection control but did not increase consistently compared to saline and needlestick control injections. This response varied by both location and proliferant. Inflammation was observed at 6 and 24 h postinjection but was resolved by 72 h compared to no-injection controls (p < 0.05). CD43+ leukocytes and ED2+ macrophages increased compared to needlestick and saline-injection control, respectively, 24 h postinjection (p < 0.05). Prolotherapy injections created an inflammatory response, but this response was variable and overall, not uniformly different from that caused by saline injections or needlestick procedures.

Immune synapse formation requires ZAP-70 recruitment by ezrin and CD43 removal by moesin.

Immunological synapse (IS) formation involves receptor-ligand pair clustering and intracellular signaling molecule recruitment with a coincident removal of other membrane proteins away from the IS. As microfilament-membrane linkage is critical to this process, we investigated the involvement of ezrin and moesin, the two ezrin/radixin/moesin proteins expressed in T cells. We demonstrate that ezrin and moesin, which are generally believed to be functionally redundant, are differentially localized and have important and complementary functions in IS formation. Specifically, we find that ezrin directly interacts with and recruits the signaling kinase ZAP-70 to the IS. Furthermore, the activation of ezrin by phosphorylation is essential for this process. In contrast, moesin dephosphorylation and removal, along with CD43, are necessary to prepare a region of the cell cortex for IS. Thus, ezrin and moesin have distinct and critical functions in the T cell cortex during IS formation.

Regulatory effects of Codonopsis lanceolata on macrophage-mediated immune responses.

Codonopsis lanceolata L. has long been used as a folk medicine in Korea, Japan and China for the treatment of lung inflammatory diseases. In this study, therefore, we aimed to demonstrate its ethnopharmacological activity by examining macrophage-function regulating effects. The total methanol extracts of fresh leaves (l-TME) or roots (r-TME) of Codonopsis lanceolata L. significantly suppressed the production of pro-inflammatory mediators (nitric oxide [NO] and tumor necrosis factor [TNF-alpha]) without altering mRNA levels. The expression of interleukin (IL)-3 and IL-6, however, was strongly diminished. According to the analysis of signaling enzyme activation by immunoblotting, phospho-IkappaB levels, a representative pro-inflammatory gene activation pathway, were not affected by the TMEs. By contrast, the Raf-ERK signaling pathway, which was involved in regulation of post-translational modification of pro-inflammatory gene products, was strongly blocked after 6-h of exposure. Moreover, l-TME down-regulated LPS-mediated phagocytic uptake and CD29-mediated cell-cell adhesion, while r-TME strongly up-regulated these two cellular events as well as fibronectin-cell adhesion. The surface levels of the costimulatory molecules (CD80 and CD86) of RAW264.7 cells were also enhanced by these extracts. l-TME also diminished functional activation (assessed by NO production) and the surface level of dectin-1, but not toll-like receptor (TLR)-2. Taken together, these data suggest that Codonopsis lanceolata may have the ability to modulate macrophage-mediated immune responses, thus contributing to its anti-inflammatory activity.