Effects of Salt Stress on Epidermal Cell Expansion in Leaves of Arabidopsis thaliana

[Objective] The aim of this study was to investigate the effects of salt stress on cell expansion in Arabidopsis thaliana rosette leaves.[Method] Arabidopsis seedlings were treated by sodium chloride at the concentration of 0,100 or 150 mmol/L. At the 7th and 14th d of treatment,with nail enamel printing mark method and computer software,the leaf blades area and abaxial epidermal pavement cells area was measured and compared using statistical analysis in Excel. [Result] The growth of Arabidopsis rosette leaves was inhibited under salt stress. Leaves treated for 7 or 14 d expanded less compared with controls. The salt-mediated decrease in leaf expansion is associated with a decrease in abaxial pavement cell expansion. [Conclusion] The decreased leaf and epidermal cell expansion under salt stress is the most important characteristic of plant physiological response to salt stress.

Effects of NaCI and Ca^(2+) on Membrane Potential of Epidermal Cells of Maize Roots

The effects of salt-stress on plants involve not only the water stress caused by low osmotic pressure, but also the toxicity of excess Na^＋. A large amount of Na^＋ entering cells would reduce K^＋ uptake, which leads to an imbalance of K：Na ratio in cells. One of the reasons for the reduced K^＋-uptake is the closure of K^＋-channel which is controlled by membrane potential. Calcium is usually applied to improve the growth of plants on saline soils and shows positive influence in the integrality of cell membrane. This study applied glass microelectrode technique to monitoring the NaCl-induced changes of membrane potential of root epidermal cells of maize （Zea mays L., Denghai 11） seedlings at NaCl concentrations of 0, 8, 20, 50, 100, 200 mmol L^-1, respectively. The effect of Ca^2＋ on the changes of membrane potential caused by NaCl was also studied. The results showed that： NaCl caused cell membrane depolarization. The depolarization became greater and faster with increasing of NaCl concentration. Moreover, the extent of depolarization was positively correlated with NaCl concentration. The addition of calcium postponed the depolarization, and decreased the degree of depolarization caused by NaCl. High NaCl concentration leads to depolarization of maize root cell membrane, which can partly be counteracted by calcium.

Ex vivo differentiation of multipotent adult progenitor cells to skin epidermal cells

Objective： By establishing the indirect contact co-culture system, we studied the in vitro condition for MAPCs differentiating into epidermal cells and the transformation of MAPCs into epidermal cell phenotype. Methods： Cell culture insert membrane was used for substitute basal membrane and MAPCs, fibroblast cells （FCs） and mixture of MAPCs and epidermal cells and FCs were separately implanted into 2 sides of it. PKH26 was used to label cloned MAPCs; type IV collagen rapid adhering method was used to isolate and culture the skin epidermal cells from l-day-old SD rat. Results： Part of the MAPCs transformed into cells expressing keratin in the presence of peripheral epithelia and FCs. Type Ⅳ collagen rapid adhering method successfully selected rats＇ epidermal stem cells. The mixture of the 2 kinds of cells or indirect culture might promote the differentiation through mesenchymal factors secreted by dermis FC. Conclusion： We were the first to have established the in vitro model of MAPCs differentiation into epidermal cells, in which MAPCs were transformed into epithelium-like cells.

Observation of two amitotic ways in the epidermal cells of plant Pelargonium zonale ‘Kleiner Liebling＇-Amitotic nuclear division for endoreduplicated cells

Amitosis, different from mitosis, is a rare form of the proliferation of the cells. Most amitosis were observed in animal cells; only few of them were reported in plant cells. The cytological investigation in this study demonstrated first time in Pelargonium zonale two different amitotic nuclear division ways： cleavage and constriction, in which no appearance of the visible chromosomes and no spindles were observed. After amitotic nuclear division the cytokinesis was also observed, in which the cytoplasm divided directly into two or more parts accompany with formation of two or more daughter cells. This study showed the genetic material may sometimes be unequally distributed between the daughter cells in amitosis, and the amitosis could lead to bi-, tri- and multinucleated cells. These new findings are discussed in regard to the nuclear amitotic process in polyploid cells which gives rise to smaller, viable nuclei in multinucleated cells with reduced numbers of genomes. It was suggested that amitosis could be a way of the division of the endoreduplicated cells.

Micromorphology of Leaf Epidermal Cells of Different Species of Cynodon dactylon in Pastoral Soil

To identify suitable grass species for growth in the Jianghan Plain,the micromorphologies of the upper and lower epidermis of the leaves of 14 bermudagrass varieties of Cynodon dactylon were investigated,and the number,length,and width of long and short cells and stomata were measured.The results showed that the anticlinal wall of epidermal cells looked like waves and the stoma was distributed in the upper and lower epidermis.However,the various indicators of the leaf epidermis of different species of Cd were significantly different.The length and width of long cells and the length of short cells of Cd Mohawk,Cd Suncity,and wild type Cd were relatively large,which may improve their ability to adapt to the pastoral soil and make them suitable for the rainy environment of the Jianghan Plain.On the contrary,the results of all indicators of Cd Xinnong 1,Cd hulled,and Cd unhulled ranked at the bottom,which was not suitable for pastoral soil habitats,while the wild type Cd could adapt to both the pastoral soil and the vegetation concrete habitat.Moreover,the long cells and short cells of Cd Kashi were tightly arranged,and the stoma in Cd Pyramid and Cd hulled were more dense,which may improve their ability to adapt to an arid environment.

Leaf Epidermal Cells： A Trap for Lipophilic Xenobiotics

Plant surfaces are covered by a layer of cuticle, which functions as a natural barrier to protect plants from mechanical damage, desiccation, and microbial invasion. Results presented in this report show that the epicuticular wax and the cuticle of plant leaves also play an important role in resisting xenobiotic invasion. Although the epicuticular wax is impermeable to hydrophilic xenobiotics, the cuticle not only restricts the penetration of hydrophilic compounds into leaf cells, but also traps lipophilic ones. The role of the epidermal cells of plant leaves in resisting xenobiotic invasion has been neglected until now. The present study shows, for the first time, that the epidermal cells may reduce or retard the transport of lipophilic xenobiotics into the internal tissues through vacuolar sequestration. Although the guard cells appear to be an easy point of entry for xenobiotics, only a very small proportion of xenobiotics present on the leaf surface actually moves into leaf tissues via the guard cells .

Enrichment of putative human epidermal stem cells based on cell size and collagen type IV adhesiveness

The enrichment and identification of human epidermal stem cells （EpSCs） are of paramount importance for both basic research and clinical application. Although several approaches for the enrichment of EpSCs have been established, enriching a pure population of viable EpSCs is still a challenging task. An improved approach is worth developing to enhance the purity and viability of EpSCs. Here we report that cell size combined with collagen type IV adhesiveness can be used in an improved approach to enrich pure and viable human EpSCs. We separated the rap- idly adherent keratinocytes into three populations that range in size from 5-7 μm （population A）, to 7-9 μm （population B）, to ≥9μm （population C） in diameter, and found that human putative EpSCs could be further enriched in population A with the smallest size. Among the three populations, population A displayed the highest density of plintegrin receptor, contained the highest percentage of cells in G0/G1 phase, showed the highest nucleus to cytoplasm ratio, and possessed the highest colony formation efficiency （CFE）. When injected into murine blastocysts, these cells participated in multi-tissue formation. More significantly, compared with a previous approach that sorted putative EpSCs according to pl-integrin antibody staining, the viability of the EpSCs enriched by the improved approach was significantly enhanced. Our results provide a putative strategy for the enrichment of human EpSCs, and encourage further study into the role of cell size in stem cell biology.

Substance P combined with epidermal stem cells promotes wound healing and nerve regeneration in diabetes mellitus

Exogenous substance P accelerates wound healing in diabetes,but the mechanism remains poorly understood.Here,we established a rat model by intraperitoneally injecting streptozotocin.Four wounds（1.8 cm diameter） were drilled using a self-made punch onto the back,bilateral to the vertebral column,and then treated using amniotic membrane with epidermal stem cells and/or substance P around and in the middle of the wounds.With the combined treatment the wound-healing rate was 100% at 14 days.With prolonged time,type I collagen content gradually increased,yet type III collagen content gradually diminished.Abundant protein gene product 9.5-and substance P-immunoreactive nerve fibers regenerated.Partial nerve fiber endings extended to the epidermis.The therapeutic effects of combined substance P and epidermal stem cells were better than with amniotic membrane and either factor alone.Our results suggest that the combination of substance P and epidermal stem cells effectively contributes to nerve regeneration and wound healing in diabetic rats.

Inhibition of epidermal growth factor receptor signaling protects human malignant glioma cells from hypoxia - induced cell death

Epidermal growth factor receptor(EGFR)signaling has become an importanttarget for drug development becauseEGFR signaling enhances tumor cell proliferation,migration,and invasion and inhibits apoptosis.However,theresults of clinical trials using EGFR inhibitors in patients with solid tumors have been disappointing.Here,wereport a protective effect of the EGFR inhibitors AG1478 and PD153035 against cell death induced by acute hy-poxia,which contrasts with their proapoptotic effects under normoxia.Under hypoxic conditions,both agents re-

Study on the Physiological,Cellular,and Morphological Aspects of the Postharvest Development of Cut Lily Flowers

It is of great practical significance to study the development process of cut lily for regulating flowering and preservation.In this study,the developmental process of lily cut flower was explored from cellular,morphological,and physiological aspects.Froma morphological aspect,the tepal edge grows faster than the midrib.The midrib groove of the flowering stage is wider than that of bud stage.The fast-growing edge of the petals results in the midrib bending outward.Moreover,the rapid growth of the stamens and stigmas also contributes to bud cracking.From the cellular aspect,in the bud stage there were more wrinkles in the outer epidermal cell wall of the tepal than those in the inner epidermal cell wall,indicating that asymmetric structural differences exist from the beginning of lily development.From a physiological aspect,from the bud cracking stage to the senescence stage,a variety of substances in the tepal cells gradually decreased,including total nitrogen,total phosphorus,total potassium,total calcium,starch,soluble sugar,and soluble protein,but not malondialdehyde.These results indicate that the asymmetric growth caused by this structural difference is responsible for flowering.The wrinkles in the cell wall can be regarded as indicators of senescent cells and are caused by the degradation of the cell wall and the loss of intracellular turgor pressure.The differences in the epidermal cells between the inner and outer tepal indicated ultrastructural changes in the tepal cells.The bud and flowering stages are maintained by the tepals acting as a sink.After flowering,the tepals gradually change from a sink to a source organ.Senescence of the cut lily flowers was caused by the decomposition of intracellular compounds in the tepals and the remobilization of nutrients from the tepals to the developing organs.

Efficacy of EGFR Tyrosine Kinase Inhibitors in Non-small Cell Lung Cancer Patients Harboring Different Types of EGFR Mutations:A Retrospective Analysis

With the development of molecular pathology, many types of epidermal growth factor receptor（EGFR） mutations have been identified. The efficacy of EGFR tyrosine kinase inhibitors（EGFR-TKIs） in non-small cell lung cancer（NSCLC） patients with different types of EGFR mutations, especially in patients with single rare mutations or complex mutations（co-occurrence of two or more different mutations）, has not been fully understood. This study aimed to examine the efficacy of EGFR-TKIs in NSCLC patients with different types of EGFR mutations. Clinical data of 809 NSCLC patients who harbored different types of EGFR mutations and treated from January 2012 to October 2016 at Renmin Hospital and Zhongnan Hospital, Wuhan, were retrospectively reviewed. The clinical characteristics of these patients and the efficacy of EGFR-TKIs were analyzed. Among these patients, 377 patients had only the EGFR del-19 mutation, 362 patients the EGFR L858R mutation in exon 21, 33 patients single rare mutations and 37 patients complex mutations. Among these 809 patients, 239 patients were treated with EGFR-TKIs. In all the 239 patients, the disease control rate（DCR） was 93.7% with two patients（0.2%） achieving complete response（CR）, the median progression free survival（PFS） was 13.0 months（95% confidence interval [CI], 11.6–14.4 months）, and the median overall survival（OS） was 55.0 months（95% CI, 26.3–83.7 months）. Subgroup analysis revealed that the DCR in patients harboring single rare or complex mutations of EGFR was significantly lower than in those with del-19 or L858 R mutation（P〈0.001）. Patients with classic mutations（del-19 and/or L858 R mutations） demonstrated longer PFS（P〈0.001） and OS（P=0.017） than those with uncommon mutations（single rare and/or complex mutations）. Furthermore, the patients with single rare mutations had shorter median OS than in those with other mutations. Multivariate Cox regression analysis identified that the type of EGFR mutations was an independent risk factor for PFS（hazard ratio [HR]=0.308, 95% CI, 0.191–0.494, P〈0.001） and OS（HR=0.221, 95% CI, 0.101–0.480, P〈0.001）. The results suggest that the single rare or complex EGFR mutations confer inferior efficacy of EGFR-TKIs treatment to the classic mutations. The prognosis of the single rare EGFR mutations is depressing. EGFR-TKIs may be not a good choice for NSCLC patients with single rare mutations of EGFR. Further studies in these patients with uncommon mutations（especially for the patients with single rare mutations） are needed to determine a better precision treatment.

Epithelial stem cell islands in the regenerated epidermis

Objective: The effects of growth factors on wound healing have been studied extensively, however, their molecular and genetic mechanisms that regulate epidermal regeneration are not fully understood. In this study, we explore the cell reversion characteristics and epithelial stem cell distribution in human regenerated epidermis treated with recombinant human epidermal growth factor (rhEGF). Methods:Tissue biospies from 8 regenerated skins treated with rhEGF were used to evaluate the cell reversion.

Threat of Domestic Power Generators to Terrestrial Ecosystem as Expressed in Some Plant Species

Polluted populations of Euphorbia heterophylla, Chromolaena odorata, Commelina diffusa and Kyllinga pumila were collected fresh from 0.1 m to 0.25 m radii of the exhausted-pipe of power generators used for domestic purposes, whereas, their non-polluted counterparts were collected from distances of 100-110 m away. The length and width of 10 leaves, each of both polluted and non-polluted populations, were measured. Their average leaf areas （LA） were 2.60 ± 0.1 cm2, 14.16 ±0.4 cm2, 5.19 ± 0.8 cm2 and 1.80 ±0.8 cm2 for the polluted populations of each of the species, respectively. Whereas, the LA of their non-polluted counterparts were 4.18±0.7 cm2, 34.39 ±0.2 cm2, 7.51± 0.1 cm2 and 10.76 ± 0.3 cm2, respectively. Damages such as plugged stomatal pores, epidermal cell aberrations and erosion, ruptured stomatal ledges, occasional leaf perforations, irregularly fused cell boundaries and glued leaf surfaces were noticeable in the leaves of the polluted populations. Such were not observed in the non-polluted populations. It was suggested that foliar morphology of these group of plants could serve as phytometer to gauge the threats of power generators used in homes to terrestrial ecosystem.

Reliable Analytic Strategy to Correlate the Morphological and Cytological Parameters on Lupinus termis L, against Fusarium oxysporum Infection

Many analytic strategies have emerged to estimate plant responses to Fusarium wilt. The demand for fast and reliable method （diagnosis, prediction） to determine isolate strength accurately is not established yet. Early determination of pathogen strength helps in plant medication. The aim of this study was to develop a faster strategy and method for early determination of fungal isolates strength in correlation to plant response. Till now, the scientists have no consensus on the most correlated parameters that could express wilt precisely. In this study, 30 isolates of Fusarium oxysporum isolated from Lupinus termis L. were used to provide an explicit image about the real strength of Fusarium isolates and its impact on the plant. Wilting percentage ranged from 26.67% to 93.33% of the infected plants depending on isolate virulence. Some of cellular, morphological and physical measurements were conducted on 8 out of 30 isolates, including root （length, fresh weight （FW） and dry weight （DW））, nodules （water content （WC）, FW, DW）, stem （height, WC, FW, DW）, total leaves/plant （WC, FW, DW） and the fourth leaf （WC, FW, DW, leaf area, epidermal cell area, epidermal cell number, succulence）. Hierarchical clustering was used to determine the variance between the isolates. Detrended correspondence analysis （DCA） and canonical correspondence analysis （CCA） were used to determine the most important growth parameters that could express wilting accurately. The CCA results showed that most of the measured parameters on the fourth leaf, except for leaf epidermal cell number, were highly and positively correlated to wilt. That makes these specific parameters valuable and sensitive for any changes in isolates strength. Accordingly, a mathematical model was created to be helpful in the quick determination of isolate strength and precise medication.

The molecular mechanisms supporting the homeostasis and activation of dendritic epidermal T cell and its role in promoting wound healing

The epidermis is the outermost layer of skin and the first barrier against invasion.Dendritic epidermal T cells(DETCs)are a subset ofT cells and an important component of the epidermal immune microenvironment.DETCs are involved in skin wound healing,malignancy and autoim-mune diseases.DETCs secrete insulin-like growth factor-1 and keratinocyte growth factor for skin homeostasis and re-epithelization and release inflammatory factors to adjust the inflammatory microenvironment of wound healing.Therefore,an understanding of their development,activation and correlative signalling pathways is indispensable for the regulation of DETCs to accelerate wound healing.Our review focuses on the above-mentioned molecular mechanisms to provide a general research framework to regulate and control the function of DETCs.

Calcium silicate accelerates cutaneous wound healing with enhanced re-epithelialization through EGF/EGFR/ERK-mediated promotion of epidermal stem cell functions

Background:Human epidermal stem cells(hESCs)play an important role in re-epithelialization and thereby in facilitating wound healing,while an effective way to activate hESCs remains to be explored.Calcium silicate(CS)is a form of bioceramic that can alter cell behavior and promote tissue regeneration.Here,we have observed the effect of CS on hESCs and investigated its possible mechanism.Methods:Using a mouse full-thickness skin excision model,we explored the therapeutic effect of CS on wound healing and re-epithelialization.In vitro,hESCs were cultured with diluted CS ion extracts(CSIEs),and the proliferation,migration ability and stemness of hESCs were evaluated.The effects of CS on the epidermal growth factor(EGF),epidermal growth factor receptor(EGFR)and extracellular signal-related kinase(ERK)signaling pathway were also explored.Results:In vivo,CS accelerated wound healing and re-epithelialization.Immunohistochemistry demonstrated that CS upregulated cytokeratin 19 and integrinβ1 expression,indicating that CS improved hESCs stemness.In vitro studies confirmed that CS improved the biological function of hESCs.And the possible mechanism could be due to the activation of the EGF/EGFR/ERK signaling pathway.Conclusion:CS can promote re-epithelialization and improve the biological functions of hESCs via activating the EGF/EGFR/ERK signaling pathway.

Spatial and Temporal Quantitative Analysis of Cell Division and Elongation Rate in Growing Wheat Leaves under Saline Conditions

Leaf growth in grasses is determined by the cell division and elongation rates, with the duration of cell elongation being one of the processes that is the most sensitive to salinity. Our objective was to investigate the distribution profiles of cell production, cell length and the duration of cell elongation in the growing zone of the wheat leaf during the steady growth phase. Plants were grown in loamy soil with or without 120 mmollL NaCI in a growth chamber, and harvested at day 3 after leaf 4 emerged. Results show that the elongation rate of leaf 4 was reduced by t20 mmollL NaCI during the steady growth phase. The distribution profile of the lengths of abaxial epidermal cells of leaf 4 during the steady growth stage shows a sigmoidal pattern along the leaf axis for both treatments. Although salinity did not affect or even increased the length of the epidermal cells in some locations in the growth zone compared to the control treatment, the final length of the epidermal cells was reduced by 14% at 120 mmollL NaCI. Thus, we concluded that the observed reduction in the leaf elongation rate derived in part from the reduced cell division rate and either the shortened cell elongation zone or shortened duration of cell elongation. This suggests that more attention should be paid to the effects of salinity on those properties of cell production and the period of cell maturation that are related to the properties of cell wall.

Genetic reporter analysis reveals an expandable reservoir of OCT4+cells in adult skin

The transcription factor Oct4(Pou5f1)is a critical regulator of pluripotency in embryonic and induced pluripotent stem cells.Therefore,Oct4 expression might identify somatic stem cell populations with inherent multipotent potential or a propensity for facilitated reprogramming.However,analysis of Oct4 expression is confounded by Oct4 pseudogenes or non-pluripotency-related isoforms.Systematic analysis of a transgenic Oct4-EGFP reporter mouse identified testis and skin as two principle sources of Oct4^(+)cells in postnatal mice.While the prevalence of GFP^(+)cells in testis rapidly declined with age,the skin-resident GFP^(+)population expanded in a cyclical fashion.These cells were identified as epidermal stem cells dwelling in the stem cell niche of the hair follicle,which endogenously expressed all principle reprogramming factors at low levels.Interestingly,skin wounding or non-traumatic hair removal robustly expanded the GFP^(+)epidermal cell pool not only locally,but also in uninjured skin areas,demonstrating the existence of a systemic response.Thus,the epithelial stem cell niche of the hair follicle harbors an expandable pool of Oct4^(+)stem cells,which might be useful for therapeutic cell transfer or facilitated reprogramming.

Petal Development in Lotus japonicus

Previous studies have demonstrated that petal shape and size in legume flowers are determined by two separate mechanisms, dorsoventral （DV） and organ internal （IN） asymmetric mechanisms, respectively. However, little is known about the molecular mechanisms controlling petal development in legumes. To address this question, we investigated petal development along the floral DV axis in Lotus japonicus with respect to cell and developmental biology by comparing wild-type legumes to mutants. Based on morphological markers, the entire course of petal development, from initiation to maturity, was grouped to define 3 phases or 13 stages. In terms of epidermal micromorphology from adaxial surface, mature petals were divided into several distinct domains, and characteristic epidermal cells of each petal differentiated at stage 9, while epidermal cells of all domains were observed until stage 12. TCP and MIXTA-like genes were found to be differentially expressed in various domains of petals at stages 9 and 12. Our results suggest that DV and IN mechanisms interplay at different stages of petal development, and their interaction at the cellular and molecular level guides the elaboration of domains within petals to achieve their ideal shape, and further suggest that TCP genes determine petal identity along the DV axis by regulatincl MIXTA-like clene expression.

Platelet-rich plasma accelerates skin wound healing by promoting re-epithelialization

Background:Autologous platelet-rich plasma(PRP)has been suggested to be effective for wound healing.However,evidence for its use in patients with acute and chronic wounds remains insufficient.The aims of this study were to comprehensively examine the effectiveness,synergy and possible mechanism of PRP-mediated improvement of acute skin wound repair.Methods:Full-thickness wounds were made on the back of C57/BL6 mice.PRP or saline solution as a control was administered to the wound area.Wound healing rate,local inflammation,angiogenesis,re-epithelialization and collagen deposition were measured at days 3,5,7 and 14 after skin injury.The biological character of epidermal stem cells(ESCs),which reflect the potential for re-epithelialization,was further evaluated in vitro and in vivo.Results:PRP strongly improved skin wound healing,which was associated with regulation of local inflammation,enhancement of angiogenesis and re-epithelialization.PRP treatment significantly reduced the production of inflammatory cytokines interleukin-17A and interleukin-1β.An increase in the local vessel intensity and enhancement of re-epithelialization were also observed in animals with PRP administration and were associated with enhanced secretion of growth factors such as vascular endothelial growth factor and insulin-like growth factor-1.Moreover,PRP treatment ameliorated the survival and activated the migration and proliferation of primary cultured ESCs,and these effects were accompanied by the differentiation of ESCs into adult cells following the changes of CD49f and keratin 10 and keratin 14.Conclusion:PRP improved skin wound healing by modulating inflammation and increasing angiogenesis and re-epithelialization.However,the underlying regulatory mechanism needs to be investigated in the future.Our data provide a preliminary theoretical foundation for the clinical administration of PRP in wound healing and skin regeneration.