Visualization of the Crossroads between a Nascent Infection Thread and the First Cell Division Event in Phaseolus vulgaris Nodulation

The development of a symbiotic nitrogen-fixing nodule in legumes involves infection and organogenesis. Infection begins when rhizobia enter a root hair through an inward structure, the infection thread (IT), which guides the bacteria towards the cortical tissue. Concurrently, organogenesis takes place by inducing cortical cell division (CCD) at the infection site. Genetic analysis showed that both events are well-coordinated; however, the dynamics connecting them remain to be elucidated. To visualize the crossroads between IT and CCD, we benefited from the fact that, in Phaseolus vulgaris nodulation, where the first division occurs in subepidermal cortical cells located underneath the infection site, we traced a Rhizobium etli strain expressing DsRed, the plant cytokinesis marker YFP-PvKNOLLE, a nuclear stain and cell wall auto-fluorescence. We found that the IT exits the root hair to penetrate an underlying subepidermal cortical (S-E) cell when it is concluding cytokinesis.     

Katanin-Dependent Microtubule Ordering in Association with ABA Is Important for Root Hydrotropism

Root hydrotropism refers to root directional growth toward soil moisture. Cortical microtubule arrays are essential for determining the growth axis of the elongating cells in plants. However, the role of microtubule reorganization in root hydrotropism remains elusive. Here, we demonstrate that the well-ordered microtubule arrays and the microtubule-severing protein KATANIN (KTN) play important roles in regulating root hydrotropism in Arabidopsis. We found that the root hydrotropic bending of the ktn1 mutant was severely attenuated but not root gravitropism. After hydrostimulation, cortical microtubule arrays in cells of the elongation zone of wild-type (WT) Col-0 roots were reoriented from transverse into an oblique array along the axis of cell elongation, whereas the microtubule arrays in the ktn1 mutant remained in disorder. Moreover, we revealed that abscisic acid (ABA) signaling enhanced the root hydrotropism of WT and partially rescued the oryzalin (a microtubule destabilizer) alterative root hydrotropism of WT but not ktn1 mutants. These results suggest that katanin-dependent microtubule ordering is required for root hydrotropism, which might work downstream of ABA signaling pathways for plant roots to search for water.     

Micro-Current Stimulation Has Potential Effects of Hair Growth-Promotion on Human Hair Follicle-Derived Papilla Cells and Animal Model

Recently, a variety of safe and effective non-pharmacological methods have been introduced as new treatments of alopecia. Micro-current electrical stimulation (MCS) is one of them. It is generally known to facilitate cell proliferation and differentiation and promote cell migration and ATP synthesis. This study aimed to investigate the hair growth-promoting effect of MCS on human hair follicle-derived papilla cells (HFDPC) and a telogenic mice model. We examined changes in cell proliferation, migration, and cell cycle progression with MCS-applied HFDPC. The changes of expression of the cell cycle regulatory proteins, molecules related to the PI3K/AKT/mTOR/Fox01 pathway and Wnt/β-catenin pathway were also examined by immunoblotting. Subsequently, we evaluated the various growth factors in developing hair follicles by RT-PCR in MCS-applied (MCS) mice model. From the results, the MCS-applied groups with specific levels showed effects on HFDPC proliferation and migration and promoted cell cycle progression and the expression of cell cycle-related proteins. Moreover, these levels significantly activated the Wnt/β-catenin pathway and PI3K/AKT/mTOR/Fox01 pathway. Various growth factors in developing hair follicles, including Wnts, FGFs, IGF-1, and VEGF-B except for VEGF-A, significantly increased in MCS-applied mice. Our results may confirm that MCS has hair growth-promoting effect on HFDPC as well as telogenic mice model, suggesting a potential treatment strategy for alopecia.     

3,4-Dehydro-L-proline Induces Programmed Cell Death in the Roots of Brachypodium distachyon

As cell wall proteins, the hydroxyproline-rich glycoproteins (HRGPs) take part in plant growth and various developmental processes. To fulfil their functions, HRGPs, extensins (EXTs) in particular, undergo the hydroxylation of proline by the prolyl-4-hydroxylases. The activity of these enzymes can be inhibited with 3,4-dehydro-L-proline (3,4-DHP), which enables its application to reveal the functions of the HRGPs. Thus, to study the involvement of HRGPs in the development of root hairs and roots, we treated seedlings of Brachypodium distachyon with 250 µM, 500 µM, and 750 µM of 3,4-DHP. The histological observations showed that the root epidermis cells and the cortex cells beneath them ruptured. The immunostaining experiments using the JIM20 antibody, which recognizes the EXT epitopes, demonstrated the higher abundance of this epitope in the control compared to the treated samples. The transmission electron microscopy analyses revealed morphological and ultrastructural features that are typical for the vacuolar-type of cell death. Using the TUNEL test (terminal deoxynucleotidyl transferase dUTP nick end labelling), we showed an increase in the number of nuclei with damaged DNA in the roots that had been treated with 3,4-DHP compared to the control. Finally, an analysis of two metacaspases’ gene activity revealed an increase in their expression in the treated roots. Altogether, our results show that inhibiting the prolyl-4-hydroxylases with 3,4-DHP results in a vacuolar-type of cell death in roots, thereby highlighting the important role of HRGPs in root hair development and root growth.     

The Effectiveness of a Pre-Exercise Performance Drink (PRX) on Indices of Maximal Cardiorespiratory Fitness

This study examined the effectiveness of a pre-exercise drink (PRX) called EM·PACT on indices of maximal cardiorespiratory fitness. Twenty-four males (n = 12) and females (n = 12) ages 18–24 years (20.25 + 1.42), volunteered as subjects. Each subject performed two randomized trials of a VO_2max treadmill test within a week of each other. Subjects in this randomized, placebo controlled, counter balanced, crossover design, ingested either a placebo (water) or PRX 20 minutes before each exercise bout. VO_2max and time to exhaustion (Time) during graded exercise testing were evaluated. Using paired samples t-tests, significantly greater mean values were found in VO_2max and Time for the PRX trial compared to the placebo trial (p < .05). Results indicate that indices of cardiorespiratory fitness; specifically VO_2max and Time are enhanced by ingestion of PRX prior to exercise testing. The combined results of this investigation may provide meaningful practical applications for coaches and athletes alike regarding ergogenic hydration options.     

Ovine Hair Follicle Stem Cells Derived from Single Vibrissae Reconstitute Haired Skin

Hair follicle stem cells (HFSCs) possess fascinating self-renewal capacity and multipotency, which play important roles in mammalian hair growth and skin wound repair. Although HFSCs from other mammalian species have been obtained, the characteristics of ovine HFSCs, as well as the methods to isolate them have not been well addressed. Here, we report an efficient strategy to obtain multipotent ovine HFSCs. Through microdissection and organ culture, we obtained keratinocytes that grew from the bulge area of vibrissa hair follicles, and even abundant keratinocytes were harvested from a single hair follicle. These bulge-derived keratinocytes are highly positive for Krt15, Krt14, Tp63, Krt19 and Itga6; in addition to their strong proliferation abilities in vitro, these keratinocytes formed new epidermis, hair follicles and sebaceous glands in skin reconstitution experiments, showing that these are HFSCs from the bulge outer root sheath. Taken together, we developed an efficient in vitro system to enrich ovine HFSCs, providing enough HFSCs for the investigations about the ovine hair cycle, aiming to promote wool production in the future.     

Kinematics Governing Mechanotransduction in the Sensory Hair of the Venus flytrap

Insects fall prey to the Venus flytrap (Dionaea muscipula) when they touch the sensory hairs located on the flytrap lobes, causing sudden trap closure. The mechanical stimulus imparted by the touch produces an electrical response in the sensory cells of the trigger hair. These cells are found in a constriction near the hair base, where a notch appears around the hair’s periphery. There are mechanosensitive ion channels (MSCs) in the sensory cells that open due to a change in membrane tension; however, the kinematics behind this process is unclear. In this study, we investigate how the stimulus acts on the sensory cells by building a multi-scale hair model, using morphometric data obtained from μ-CT scans. We simulated a single-touch stimulus and evaluated the resulting cell wall stretch. Interestingly, the model showed that high stretch values are diverted away from the notch periphery and, instead, localized in the interior regions of the cell wall. We repeated our simulations for different cell shape variants to elucidate how the morphology influences the location of these high-stretch regions. Our results suggest that there is likely a higher mechanotransduction activity in these ’hotspots’, which may provide new insights into the arrangement and functioning of MSCs in the flytrap.Dataset: 10.3929/ethz-b-000448954     

Root growth and phosphorus and potassium uptake by two corn genotypes in the field

In a field study P and K uptake by two corn (Zea mays L.) genotypes which differed in root growth was investigated. The effect of differences in root growth on P and K uptake was assessed using a mechanistic-mathematical model which describes nutrient uptake by growing plant roots in soil. Nitrogen was applied at 0 and 227 kg ha^–1 to Pioneer 3732 and B73xMo17 corn grown on Raub silt loam (fine-silty, mixed, mesic Aquic Argiudoll) and at 227 kg N ha^–1 to these two genotypes on Chalmers silt loam (finesilty, mixed, mesic, Typic Haplaquoll). Root growth and P and K uptake by the two corn genotypes was measured 31, 47, 75 and 91 d after planting on the Raub and 31, 47, 61 and 75 d after planting on the Chalmers soil.Root growth and P and K uptake by B73xMo17 was greater than that of Pioneer 3732 on N-fertilized Raub soil. On Chalmers soil the difference in root growth between the two genotypes resulted in an increase in K but not P uptake. The higher soil P level of the Chalmers appears to have offset possible differences in P uptake due to root size. There were no differences between the two genotypes in either the percentage of roots with root hairs, or the density or length of root hairs. Phosphorus and K uptake calculated with the simulation models for both corn genotypes on both soils over each of three growth periods agreed with observed P (Y = 0.68X + 1.71; r = 0.944^**) and K (Y = 0.88X + 15.52; r = 0.928^**) uptake. Differences in P and K uptake between B73xMo17 and Pioneer 3732 resulted primarily from the difference in root growth in the topsoil. A high correlation was found between root surface area and P (r = 0.893^**) and K (r = 0.928^**) uptake by both corn genotypes on both the soils.Journal paper No. 10,316 Purdue Univ. Agric. Exp. Stn., W. Lafayette, In 47906. Contribution from the Dep. of Agron. This paper was supported in part by a grant from the Tennessee Valley Authority.     

Regulation of Cell Division and Growth in Roots of <Emphasis Type="Italic">Lactuca sativa</Emphasis> L. Seedlings by the <Emphasis Type="Italic">Ent</Emphasis>-Kaurene Diterpenoid Rabdosin B

Rabdosin B, an ent-kaurene diterpenoid purified from the air-dried aerial parts of Isodon japonica (Burm.f) Hara var. galaucocalyx (maxin) Hara, showed a biphasic, dose-dependent effect on root growth and a strong inhibitory effect on root hair development in lettuce seedlings (Lactuca sativa L.). Lower concentrations of rabdosin B (20–80 μM) significantly promoted root growth, but its higher levels at 120–200 μM, by contrast, had inhibitory effects. Additionally, all tested concentrations (10–40 μM) inhibited root hair development of seedlings in a dose-dependent manner. Further investigations on the underlying mechanism revealed that the promotion effect of rabdosin B at the lower concentrations resulted from increasing the cell length in the mature region and enhancing the mitotic activity of meristematic cells in seedlings’ root tips. In contrast, rabdosin B at higher concentrations inhibited root growth by affecting both cell length in the mature region and division of meristematic cells. Comet assay and cell cycle analysis demonstrated that the decrease of mitotic activity of root meristematic cells was due to DNA damage induced cell cycle retardation of the G₂ phase and S phase at different times.     

Alfalfa Leaf Saponins and Insect Resistance

Dried alfalfa (Medicago sativa) leaf tissue incorporated in artificial diet to give a final concentration of 0.5 or 1.6 mg/g fresh weight of saponins significantly inhibited growth and development of larvae of the European corn borer (Ostrinia nubilalis). Saponin fractions isolated from root or shoot tissues of alfalfa also inhibited growth when incorporated at equivalent concentrations but had little effect on development. Root saponins were somewhat more harmful than shoot saponins.     

Sound Waves Promote Arabidopsis thaliana Root Growth by Regulating Root Phytohormone Content

Sound waves affect plants at the biochemical, physical, and genetic levels. However, the mechanisms by which plants respond to sound waves are largely unknown. Therefore, the aim of this study was to examine the effect of sound waves on Arabidopsis thaliana growth. The results of the study showed that Arabidopsis seeds exposed to sound waves (100 and 100 + 9k Hz) for 15 h per day for 3 day had significantly longer root growth than that in the control group. The root length and cell number in the root apical meristem were significantly affected by sound waves. Furthermore, genes involved in cell division were upregulated in seedlings exposed to sound waves. Root development was affected by the concentration and activity of some phytohormones, including cytokinin and auxin. Analysis of the expression levels of genes regulating cytokinin and auxin biosynthesis and signaling showed that cytokinin and ethylene signaling genes were downregulated, while auxin signaling and biosynthesis genes were upregulated in Arabidopsis exposed to sound waves. Additionally, the cytokinin and auxin concentrations of the roots of Arabidopsis plants increased and decreased, respectively, after exposure to sound waves. Our findings suggest that sound waves are potential agricultural tools for improving crop growth performance.     

Identification and Quantification of the Main Active Anticancer Alkaloids from the Root of Glaucium flavum

Glaucium flavum is used in Algerian folk medicine to remove warts (benign tumors). Its local appellations are Cheqiq el-asfar and Qarn el-djedyane. We have recently reported the anti-tumoral activity of Glaucium flavum root alkaloid extract against human cancer cells, in vitro and in vivo. The principal identified alkaloid in the extract was protopine. This study aims to determine which component(s) of Glaucium flavum root extract might possess potent antitumor activity on human cancer cells. Quantitative estimation of Glaucium flavum alkaloids was realized by HPLC-DAD. Glaucium flavum effect on human normal and cancer cell viability was determined using WST-1 assay. Quantification of alkaloids in Glaucium flavum revealed that the dried root part contained 0.84% of protopine and 0.07% of bocconoline (w/w), while the dried aerial part contained only 0.08% of protopine, glaucine as the main alkaloid, and no bocconoline. In vitro evaluation of the growth inhibitory activity on breast cancer and normal cells demonstrated that purified protopine did not reproduce the full cytotoxic activity of the alkaloid root extract on cancer cell lines. On the other hand, bocconoline inhibited strongly the viability of cancer cells with an IC₅₀ of 7.8 μM and only a low cytotoxic effect was observed against normal human cells. Our results showed for the first time that protopine is the major root alkaloid of Glaucium flavum. Finally, we are the first to demonstrate a specific anticancer effect of Glaucium flavum root extract against breast cancer cells, which can be attributed, at least in part, to bocconoline.     

Phytotoxic Effects of Leukamenin E (an ent-kaurene diterpenoid) on Root Growth and Root Hair Development in Lactuca sativa L. seedlings

Leukamenin E, an ent-kaurene diterpenoid isolated from Isodon racemosa (Hemsl) Hara, showed phytotoxic effects on root growth and root hair development of lettuce seedlings (Lactuca sativa L.). Lower concentrations (10 μM) of leukamenin E did not affect root growth, but at concentrations higher than 50 μM, the rate was inhibited. The influence of leukamenin E on root growth rate was closely correlated with alterations in the mitotic index. A low incidence of aberrant mitosis image was observed when lettuce roots were treated with higher concentrations (100 and 200 μM) of leukamenin E. This suggests that inhibition of root growth may be due to inhibition of cell division. All tested concentrations of the diterpenoid (10 μM or more) inhibited root hair development in a dose-dependent manner. At a concentration of 80 μM, leukamenin E completely blocked root hair initiation. Application of Ag⁺—an ethylene action inhibitor—to lettuce seedlings inhibited root hair elongation similar to the diterpenoid. Enhanced root hair length was stimulated by exogenous ethephon—an ethylene-releasing agent—and could be reversed by addition of leukamenin E. This suggests that leukamenin E may act as a potential ethylene action antagonist in the inhibition of lettuce root hair development. We conclude that leukamenin E may curb root hair development by interfering with ethylene action at concentrations above 10 μM and inhibits root growth via inhibition of cell division at concentrations above 50 μM. Lan Ding and Linlin Qi contributed equally to this work.     

Effects of Allelochemical Stress Produced by Sicyos deppei on Seedling Root Ultrastructure of Phaseolus vulgaris and Cucurbita ficifolia

The allelopathic potential of the cucurbit Sicyos deppei has been reported. Aqueous leachates (1% w/v) of S. deppei significantly inhibited the radicle growth of several test plants. Root tips of treated plants were thicker, with a brownish color as compared to control roots. Light microscope observations of treated root tips of Phaseolus vulgaris showed that cells around the quiescent center appeared compressed. Tissues were disorganized and lacked evident cell differentiation. Cucurbita ficifolia did not show these differences in root morphology. Scanning and electron microscope observations showed that treated roots of both P. vulgaris and C. ficifolia had ultrastructural alterations of certain organelles, the plasma membrane, and cell walls. Root cap cells of treated roots showed amorphous and inactivated (nondividing) nuclei, mitochondria, and endoplasmic reticulum. Vacuoles in treated cells showed invaginations that may correspond to engulfing of damaged cellular components.     

Possible Role of Circulating Bone Marrow Mesenchymal Progenitors in Modulating Inflammation and Promoting Wound Repair

Circulating bone marrow mesenchymal progenitors (BMMPs) are known to be potent antigen-presenting cells that migrate to damaged tissue to secrete cytokines and growth factors. An altered or dysregulated inflammatory cascade leads to a poor healing outcome. A skin model developed in our previous study was used to observe the immuno-modulatory properties of circulating BMMP cells in inflammatory chronic wounds in a scenario of low skin perfusion. BMMPs were analysed exclusively and in conjunction with recombinant tumour necrosis factor alpha (TNFα) and recombinant hepatocyte growth factor (HGF) supplementation. We analysed the expression levels of interleukin-8 (IL-8) and ecto-5′-nucleotidase (CD73), together with protein levels for IL-8, stem cell factor (SCF), and fibroblast growth factor 1 (FGF-1). The successfully isolated BMMPs were positive for both hemopoietic and mesenchymal markers and showed the ability to differentiate into adipocytes, chondrocytes, and osteocytes. Significant differences were found in IL-8 and CD73 expressions and IL-8 and SCF concentrations, for all conditions studied over the three time points taken into consideration. Our data suggests that BMMPs may modulate the inflammatory response by regulating IL-8 and CD73 and influencing IL-8 and SCF protein secretions. In conclusion, we suggest that BMMPs play a role in wound repair and that their induced application might be suitable for scenarios with a low skin perfusion.