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OBJECTIVE:Epidermal stem cells (ESCs) play a critical role in wound healing, but the mechanism underlying ESC proliferation is not well defined. Here, we explore the effects of RhoA on ESC proliferation and the possible underlying mechanism. METHODS:Human ESCs were enriched by rapid adhesion to collagen IV. RhoA(+/+)(G14V), RhoA(-/-)(T19N) and pGFP control plasmids were transfected into human ESCs. The effect of RhoA on cell proliferation was detected by cell proliferation and DNA synthesis assays. Induction of PKN1 activity by RhoA was determined by immunoblot analysis, and the effects of PKN1 on RhoA in terms of inducing cell proliferation and cyclin D1 expression were detected using specific siRNA targeting PKN1. The effects of U-46619 (a RhoA agonist) and C3 transferase (a RhoA antagonist) on ESC proliferation were observed in vivo. RESULTS:RhoA had a positive effect on ESC proliferation, and PKN1 activity was up-regulated by the active RhoA mutant (G14V) and suppressed by RhoA T19N. Moreover, the ability of RhoA to promote ESC proliferation and DNA synthesis was interrupted by PKN1 siRNA. Additionally, cyclin D1 protein and mRNA expression levels were up-regulated by RhoA G14V, and these effects were inhibited by siRNA-mediated knock-down of PKN1. RhoA also promoted ESC proliferation via PKN in vivo. CONCLUSION:This study shows that the effect of RhoA on ESC proliferation is mediated by activation of the PKN1-cyclin D1 pathway in vitro, suggesting that RhoA may serve as a new therapeutic target for wound healing.

Mesenchymal stem cells (MSCs) are promising candidates for skin wound repair due to their capabilities of accumulating at wounds and differentiating into multiple types of skin cells. However, the underlying mechanisms responsible for these processes remain unclear. In this study, we found that osteopontin (OPN) stimulated the migration of MSCs in vitro, and observed the recruitment of endogenous MSCs to a skin wound and their differentiation into keratinocytes and endothelial cells. In OPN knock-out mice, the recruitment of MSCs to the skin wound was significantly inhibited, and wound closure was hampered after an intradermal injection of exogenous MSCs compared to wild-type mice. Consistent with these observations, the expressions of adhesion molecule CD44 and its receptor E-selectin were significantly decreased in the lesions of OPN knock-out mice compared with wild-type mice suggesting that OPN may regulate the migration of MSCs through its interactions with CD44 during skin wound recovery. In summary, our data demonstrated that OPN played a critical role in activating the migration of MSCs to injured sites and their differentiation into specific skin cell types during skin wound healing.

Hypertrophic scars (HS) are skin disorders which occur after wounding and thermal injury. Our previous studies have suggested that secreted frizzled-related protein 2 (SFRP2) is involved in HS formation and that the suppression of SFRP2 promotes apoptosis of hypertrophic scar fibroblasts (HSFBs). However, the mechanisms have not been clarified. Previous studies revealed that Slug expression inhibits cell apoptosis, in vitro and in vivo, and SFRP2 regulates the expression of Slug in cervical cancer cells. In the present study, we quantified differential expression levels of expression of SFRP2 and Slug in HS and normal skin tissues by immunohistochemistry, both of which have important anti-apoptosis roles. Furthermore, a short hairpin RNA approach was adopted to investigate the potential function of SFRP2 and Slug in HSFB apoptosis. Cell apoptosis was detected using fluorescence-activated cell sorting and Caspase-3 activity was assayed by spectrophotometry. This study demonstrates that SFRP2 expression, as well as Slug, is dramatically up-regulated in HS relative to normal skin tissues, and the Slug expression is positively correlated with SFRP2. Slug expression was down-regulated in SFRP2-deficient cells, and the down-regulation of Slug expression increased sensitivity to apoptosis which was induced through a caspase-3-dependent pathway. The infected cells with reduced levels of Slug were tested for the expression of apoptosis-related genes (Bcl-2, Bax and PUMA) which were previously identified as Slug targets. Bcl-2 expression was down-regulated in Slug-deficient cells. In conclusion, SFRP2 appears to interact with Slug to affect the apoptosis of hypertrophic scar fibroblasts.

The mechanisms of hypertrophic scar formation are not fully understood. We previously screened the differentially expressed genes of human hypertrophic scar tissue and identified P311 gene as upregulated. As the activities of P311 in human fibroblast function are unknown, we examined the distribution of it and the effects of forced expression or silencing of expression of P311. P311 expression was detected in fibroblast-like cells from the hypertrophic scar of burn injury patients but not in peripheral blood mononuclear cells, bone marrow mesenchymal stem cells, epidermal cells or normal skin dermal cells. Transfection of fibroblasts with P311 gene stimulated the expression of alpha-smooth muscle actin (alpha-SMA), TGF-beta1 and alpha1(I) collagen (COL1A1), and enhanced the contraction of fibroblast populated collagen lattices (FPCL). In contrast, interference of fibroblast P311 gene expression decreased the TGF-beta1 mRNA expression and reduced the contraction of fibroblasts in FPCL. These results suggest that P311 may be involved in the pathogenesis of hypertrophic scar via induction of a myofibroblastic phenotype and of functions such as TGF-beta1 expression. P311 could be a novel target for the control of hypertrophic scar development.

Photosynthetic-nitrogen use efficiency (PNUE) is a useful trait to characterize leaf physiology and survival strategy. PNUE can also be considered as part of 'leaf economics spectrum' interrelated with leaf nutrient concentrations, photosynthesis and respiration, leaf life-span and dry-mass investment. However, few studies have paid attention to PNUE of N-fixing tree seedlings in subtropical China. In this study, we investigated the differences in PNUE, leaf nitrogen (N) allocation, and mesophyll conductance (gm) in Dalbergia odorifera and Erythrophleum fordii (N-fixing trees), and Betula alnoides and Castanopsis hystrix (non-N-fixing trees). PNUE of D. odorifera and E. fordii were significantly lower than those of B. alnoides and C. hystrix mainly because of their allocation of a lower fraction of leaf N to Rubisco (PR) and bioenergetics (PB). Mesophyll conductance had a significant positive correlation with PNUE in D. odorifera, E. fordii, and B. alnoides, but the effect of gm on PNUE was different between species. The fraction of leaf N to cell wall (PCW) had a significant negative correlation with PR in B. alnoides and C. hystrix seedling leaves, but no correlation in D. odorifera and E. fordii seedling leaves, which may indicate that B. alnoides and C. hystrix seedling leaves did not have enough N to satisfy the demand from both the cell wall and Rubisco. Our results indicate that B. alnoides and C. hystrix may have a higher competitive ability in natural ecosystems with fertile soil, and D. odorifera and E. fordii may grow well in N-poor soil. Mixing these non-N-fixing and N-fixing trees for afforestation is useful for improving soil N utilization efficiency in the tropical forests.

Variation in photosynthetic-nitrogen use efficiency (PNUE) is generally affected by several factors such as leaf nitrogen allocation and leaf diffusional conductances to CO2, although it is still unclear which factors significantly affect PNUE in tropical montane rain forest trees. In this study, comparison of PNUE, photosynthetic capacity, leaf nitrogen allocation, and diffusional conductances to CO2 between five Fagaceae tree species and five Leguminosae tree species were analyzed in Jianfengling tropical montane rain forest, Hainan Island, China. The result showed that PNUE of Fagaceae was significantly higher than that of Leguminosae (+35.5%), attributed to lower leaf nitrogen content per area (Narea, -29.4%). The difference in nitrogen allocation was the main biochemical factor that influenced interspecific variation in PNUE of these tree species. Fagaceae species allocated a higher fraction of leaf nitrogen to the photosynthetic apparatus (PP, +43.8%), especially to Rubisco (PR, +50.0%) and bioenergetics (PB +33.3%) in comparison with Leguminosae species. Leaf mass per area (LMA) of Leguminosae species was lower than that of Fagaceae species (-15.4%). While there was no significant difference shown for mesophyll conductance (gm), Fagaceae tree species may have greater chloroplast to total leaf surface area ratios and that offset the action of thicker cell walls on gm. Furthermore, weak negative relationship between nitrogen allocation in cell walls and in Rubisco was found for Castanopsis hystrix, Cyclobalanopsis phanera and Cy. patelliformis, which might imply that nitrogen in the leaves was insufficient for both Rubisco and cell walls. In summary, our study concluded that higher PNUE might contribute to the dominance of most Fagaceae tree species in Jianfengling tropical montane rain forest.

Ensemble forecasting is advocated as a way of reducing uncertainty in species distribution modeling (SDM). This is because it is expected to balance accuracy and robustness of SDM models. However, there are little available data regarding the spatial similarity of the combined distribution maps generated by different consensus approaches. Here, using eight niche-based models, nine split-sample calibration bouts (or nine random model-training subsets), and nine climate change scenarios, the distributions of 32 forest tree species in China were simulated under current and future climate conditions. The forecasting ensembles were combined to determine final consensual prediction maps for target species using three simple consensus approaches (average, frequency, and median [PCA]). Species' geographic ranges changed (area change and shifting distance) in response to climate change, but the three consensual projections did not differ significantly with respect to how much or in which direction, but they did differ with respect to the spatial similarity of the three consensual predictions. Incongruent areas were observed primarily at the edges of species' ranges. Multiple stepwise regression models showed the three factors (niche marginality and specialization, and niche model accuracy) to be related to the observed variations in consensual prediction maps among consensus approaches. Spatial correspondence among prediction maps was the highest when niche model accuracy was high and marginality and specialization were low. The difference in spatial predictions suggested that more attention should be paid to the range of spatial uncertainty before any decisions regarding specialist species can be made based on map outputs. The niche properties and single-model predictive performance provide promising insights that may further understanding of uncertainties in SDM.

Diffusional limitations to photosynthesis, relative water content (RWC), pigment concentrations and their association with reflectance indices were studied in olive (Olea europaea) saplings subjected to water-stress and re-watering. RWC decreased sharply as drought progressed. Following rewatering, RWC gradually increased to pre-stress values. Photosynthesis (A), stomatal conductance (gs), mesophyll conductance (gm), total conductance (gt), photochemical reflectance index (PRI), water index (WI) and relative depth index (RDI) closely followed RWC. In contrast, carotenoid concentration, the carotenoid to chlorophyll ratio, water content reflectance index (WCRI) and structural independent pigment index (SIPI) showed an opposite trend to that of RWC. Photosynthesis scaled linearly with leaf conductance to CO2; however, A measured under non-photorespiratory conditions (A1%O2) was approximately two times greater than A measured at 21% [O2], indicating that photorespiration likely increased in response to drought. A1%O2 also significantly correlated with leaf conductance parameters. These relationships were apparent in saturation type curves, indicating that under non-photorespiratory conditions, CO2 conductance was not the major limitations to A. PRI was significant correlated with RWC. PRI was also very sensitive to pigment concentrations and photosynthesis, and significantly tracked all CO2 conductance parameters. WI, RDI and WCRI were all significantly correlated with RWC, and most notably to leaf transpiration. Overall, PRI correlated more closely with carotenoid concentration than SIPI; whereas WI tracked leaf transpiration more effectively than RDI and WCRI. This study clearly demonstrates that PRI and WI can be used for the fast detection of physiological traits of olive trees subjected to water-stress.

It is inevitable that tree species will undergo considerable range shifts in response to anthropogenic induced climate change, even in the near future. Species Distribution Models (SDMs) are valuable tools in exploring general temporal trends and spatial patterns of potential range shifts. Understanding projections to future climate for tree species will facilitate policy making in forestry. Comparative studies for a large number of tree species require the availability of suitable and standardized indices. A crucial limitation when deriving such indices is the threshold problem in defining ranges, which has made interspecies comparison problematic until now. Here we propose a set of threshold-free indices, which measure range explosion (I), overlapping (O), and range center movement in three dimensions (Dx, Dy, Dz), based on fuzzy set theory (Fuzzy Set based Potential Range Shift Index, F-PRS Index). A graphical tool (PRS_Chart) was developed to visualize these indices. This technique was then applied to 46 Pinaceae species that are widely distributed and partly common in China. The spatial patterns of the modeling results were then statistically tested for significance. Results showed that range overlap was generally low; no trends in range size changes and longitudinal movements could be found, but northward and poleward movement trends were highly significant. Although range shifts seemed to exhibit huge interspecies variation, they were very consistent for certain climate change scenarios. Comparing the IPCC scenarios, we found that scenario A1B would lead to a larger extent of range shifts (less overlapping and more latitudinal movement) than the A2 and the B1 scenarios. It is expected that the newly developed standardized indices and the respective graphical tool will facilitate studies on PRS's for other tree species groups that are important in forestry as well, and thus support climate adaptive forest management.

PURPOSE: The aim of this study is to determine the predictive value of preoperative blood neutrophil-to-lymphocyte ratio (NLR) for recurrence in recurrent hepatocellular carcinoma (RHCC) patients following thermal ablation. MATERIAL AND METHODS: This retrospective study enrolled 506 RHCC patients who underwent thermal ablation from April 2006 to April 2014. The clinicopathological parameters including NLR were evaluated to identify predictors of recurrence rate after thermal ablation. A Cox multiple regression analysis was performed to determine the parameters that predicted recurrence in RHCC patients. The best cutoff value of NLR was determined with time-dependent receiver operating characteristic (ROC) curve analysis. The recurrence-free survival (RFS) rate was determined in patients with high and low NLR. RESULTS: The multivariate Cox proportional hazard model analysis showed that NLR was a prognostic factor in recurrence-free survival. NLR ≥ 2.14 was evaluated (AUROC = 0.824; P

A range of factors has been identified that affect the temperature sensitivity (Q₁₀ values) of the soil-to-atmosphere CO₂ flux. However, the factors influencing the spatial distribution of Q₁₀ values within warm temperate forests are poorly understood. In this study, we examined the spatial variation of Q₁₀ values and its controlling factors in both a naturally regenerated oak forest (OF) and a pine plantation (PP). Q₁₀ values were determined based on monthly soil respiration (R(S)) measurements at 35 subplots for each stand from Oct. 2008 to Oct. 2009. Large spatial variation of Q₁₀ values was found in both OF and PP, with their respective ranges from 1.7 to 5.12 and from 2.3 to 6.21. In PP, fine root biomass (FR) (R = 0.50, P = 0.002), non-capillary porosity (NCP) (R = 0.37, P = 0.03), and the coefficients of variation of soil temperature at 5 cm depth (CV of T₅) (R = -0.43, P = 0.01) well explained the spatial variance of Q₁₀. In OF, carbon pool lability reflected by light fractionation method (LLFOC ) well explained the spatial variance of Q₁₀ (R = -0.35, P = 0.04). Regardless of forest type, LLFOC and FR correlation with the Q₁₀ values were significant and marginally significant, respectively; suggesting a positive relationship between substrate availability and apparent Q₁₀ values. Parameters related to gas diffusion, such as average soil water content (SWC) and NCP, negatively or positively explained the spatial variance of Q₁₀ values. Additionally, we observed significantly higher apparent Q₁₀ values in PP compared to OF, which might be partly attributed to the difference in soil moisture condition and diffusion ability, rather than different substrate availabilities between forests. Our results suggested that both soil chemical and physical characters contributed to the observed large Q₁₀ value variation.

Predicting species range shifts in response to climatic change is a central aspect of global change studies. An ever growing number of species have been modeled using a variety of species distribution models (SDMs). However, quantitative studies of the characteristics of range shifts are rare, predictions of range changes are hard to interpret, analyze and summarize, and comparisons between the various models are difficult to make when the number of species modeled is large. Maxent was used to model the distribution of 12 Abies spp. in China under current and possible future climate conditions. Two fuzzy set defined indices, range increment index (I) and range overlapping index (O), were used to quantify range shifts of the chosen species. Correlation analyses were used to test the relationships between these indices and species distribution characteristics. Our results show that Abies spp. range increments (I) were highly correlated with longitude, latitude, and mean roughness of their current distributions. Species overlapping (O) was moderately, or not, correlated with these parameters. Neither range increments nor overlapping showed any correlation with species prevalence. These fuzzy sets defined indices provide ideal measures of species range shifts because they are stable and threshold-free. They are reliable indices that allow large numbers of species to be described, modeled, and compared on a variety of taxonomic levels.