BackgroundBirch bark has a long lasting history as a traditional medicinal remedy to accelerate wound healing. Recently, the efficacy of birch bark preparations has also been proven clinically. As active principle pentacyclic triterpenes are generally accepted. Here, we report a comprehensive study on the underlying molecular mechanisms of the wound healing properties of a well-defined birch bark preparation named as TE (triterpene extract) as well as the isolated single triterpenes in human primary keratinocytes and porcine ex-vivo wound healing models.Methodology/Principal FindingsWe show positive wound healing effects of TE and betulin in scratch assay experiments with primary human keratinocytes and in a porcine ex-vivo wound healing model (WHM). Mechanistical studies elucidate that TE and betulin transiently upregulate pro-inflammatory cytokines, chemokines and cyclooxygenase-2 on gene and protein level. For COX-2 and IL-6 this increase of mRNA is due to an mRNA stabilizing effect of TE and betulin, a process in which p38 MAPK and HuR are involved. TE promotes keratinocyte migration, putatively by increasing the formation of actin filopodia, lamellipodia and stress fibers. Detailed analyses show that the TE components betulin, lupeol and erythrodiol exert this effect even in nanomolar concentrations. Targeting the actin cytoskeleton is dependent on the activation of Rho GTPases.Conclusion/SignificanceOur results provide insights to understand the molecular mechanism of the clinically proven wound healing effect of birch bark. TE and betulin address the inflammatory phase of wound healing by transient up-regulation of several pro-inflammatory mediators. Further, they enhance migration of keratinocytes, which is essential in the second phase of wound healing. Our results, together with the clinically proven efficacy, identify birch bark as the first medical plant with a high potential to improve wound healing, a field which urgently needs effective remedies.

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BackgroundCytokines such as TNF-alpha and IL-1beta are known for their contribution to inflammatory processes in liver. In contrast, the cytokine IL-17 has not yet been assigned a role in liver diseases. IL-17 can cooperate with TNF-alpha to induce a synergistic response on several target genes in different cell lines, but no data exist for primary hepatocytes. To enhance our knowledge on the impact of IL-17 alone and combined with TNF-alpha in primary murine hepatocytes a comprehensive microarray study was designed. IL-1beta was included as this cytokine is suggested to act in a similar manner as the combination of TNF-alpha and IL-17, especially with respect to its role in mRNA stabilization.ResultsThe present microarray analysis demonstrates that primary murine hepatocytes responded to IL-17 stimulation by upregulation of chemokines and genes, which are functionally responsible to increase and sustain inflammation. Cxcl2, Nfkbiz and Zc3h12a were strongly induced, whereas the majority of the genes were only very moderately up-regulated. Promoter analysis revealed involvement of NF-kappaB in the activation of many genes. Combined stimulation of TNF-alpha/IL-17 resulted in enhanced induction of gene expression, but significantly synergistic effects could be applied only to a few genes, such as Nfkbiz, Cxcl2, Zc3h12 and Steap4. Comparison of the gene expression profile obtained after stimulation of TNF-alpha/IL-17 versus IL-1beta proposed an "IL-1beta-like effect" of the latter cytokine combination. Moreover, evidence was provided that modulation of mRNA stability may be a major mechanism by which IL-17 regulates gene expression in primary hepatocytes. This assumption was exemplarily proven for Nfkbiz mRNA for the first time in hepatocytes. Our studies also suggest that RNA stability can partially be correlated to the existence of AU rich elements, but further mechanisms like the RNase activity of the up-regulated Zc3h12a have to be considered.ConclusionsOur microarray analysis gives new insights in IL-17 induced gene expression in primary hepatocytes highlighting the crosstalk with the NF-kappaB signaling pathway. Gene expression profile suggests IL-17 alone and in concert with TNF-alpha a role in sustaining liver inflammatory processes. IL-17 might exceed this function by RNA stabilization.

Pancreatic cancer progresses aggressively and owing to chemoresistance responds poorly to chemotherapy. Thus, there is an urgent need to understand the mechanisms of cancer cell resistance to generate effective strategies to circumvent intrinsic chemoresistance in this tumour indication. In this study, three pancreatic cancer cell lines, MIA PaCa-2, MDAPanc-3 and AsPC-1, were treated with the proteasome inhibitor MG-132 together with camptothecin, doxorubicin or paclitaxel, and cytotoxicity was measured using the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay. The combination of MG-132 and camptothecin at a ratio of 5 : 1 gave the most promising results and enhanced cytotoxicity compared with the single compounds in MIA PaCa-2 cells. The increase was shown to be due to enhanced caspase-3 activity resulting in apoptosis. Moreover, this combination upregulated the levels of the proapoptotic protein Noxa and reduced the levels of the antiapoptotic protein Mcl-1, as demonstrated by western blotting. In contrast, the combination of MG-132 with doxorubicin also induced increased cytotoxicity, but apoptosis was decreased. The lack of an enhanced apoptosis induction could be correlated with high levels of Mcl-1 in response to the combined treatment with MG-132 and doxorubicin. Thus, the results indicate that regulation of the antiapoptotic and proapoptotic Bcl-2 family members Noxa and Mcl-1 is predicative of the effectiveness of the combination of MG-132 with different anticancer agents on apoptosis induction in pancreatic cancer cells.

<p><strong>Abstract.</strong> We evaluated how ranges of four endemic and non-endemic aquatic ostracode species changed in response to long-term (glacial-interglacial cycles) and abrupt climate fluctuations during the last 155 ka in the northern Neotropical region. We employed two complementary approaches, fossil records and species distribution modeling (SDM). Fossil assemblages were obtained from sediment cores PI-1, PI-2, PI-6 and Petén-Itzá 22-VIII-99 from Petén Itzá Scientific Drilling Project, Lake Petén Itzá, Guatemala. To obtain a spatially resolved pattern of (past) species distribution, a downscaling cascade is employed. SDM's were reconstructed for the Last Interglacial (~ 120 ka BP), the Last Glacial Maximum (~ 22 ka BP) and the middle Holocene (~ 6 ka BP). During glacial/interglacial cycles and Marine Isotope Stages, modeled paleo-distributions and paleo-records show nearly continuous presence of endemic and non-endemic species in the region, suggesting negligible effects of long-term climate variations on aquatic niche stability. During periods of abrupt ecological disruption such as Heinrich Stadial 1 (HS1), endemic species were resilient, remaining within their current areas of distribution. Non-endemic species, however, proved to be more sensitive. Modeled paleo-distributions suggest that the geographic range of non-endemic species changed, moving southward into Central America. Due to the uncertainties involved in the downscaling from the global numerical to the highly resolved regional geospatial statistical modelling, results can be seen as benchmark for future studies using similar approaches. Given relatively moderate temperature decreases in Lake Petén Itzá waters (~ 5 ºC) and persistence of some aquatic ecosystems even during periods of severe drying in HS1, our data suggest 1) existence of micro-refugia and/or 2) continuous interaction between central metapopulations and surrounding populations, enabling aquatic taxa to survive climate fluctuations in the northern Neotropical region.</p>