Deep simple epicotyl morphophysiological dormancy in seeds of two Viburnum species, with special reference to shoot growth and development inside the seed

Background and Aims

In seeds with deep simple epicotyl morphophysiological dormancy, warm and cold stratification are required to break dormancy of the radicle and shoot, respectively. Although the shoot remains inside the seed all winter, little is known about its growth and morphological development prior to emergence in spring. The aims of the present study were to determine the temperature requirements for radicle and shoot emergence in seeds of Viburnum betulifolium and V. parvifolium and to monitor growth of the epicotyl, plumule and cotyledons in root-emerged seeds.

Methods

Fresh and pre-treated seeds of V. betulifolium and V. parvifolium were incubated under various temperature regimes and monitored for radicle and shoot emergence. Growth of the epicotyl and cotyledons at different stages was observed with dissecting and scanning electron microscopes.

Key Results

The optimum temperature for radicle emergence of seeds of both species, either kept continuously at a single regime or exposed to a sequence of regimes, was 20/10 °C. GA₃ had no effect on radicle emergence. Cold stratification (5 °C) was required for shoot emergence. The shoot apical meristem in fresh seeds did not form a bulge until the embryo had grown to the critical length for radicle emergence. After radicle emergence, the epicotyl–plumule and cotyledons grew slowly at 5 and 20/10 °C, and the first pair of true leaves was initiated. However, the shoot emerged only from seeds that received cold stratification.

Conclusions

Seeds of V. betulifolium and V. parvifolium have deep simple epicotyl morphophysiological dormancy, C_1bB (root)–C₃ (epicotyl). Warm stratification was required to break the first part of physiological dormancy (PD), thereby allowing embryo growth and subsequently radicle emergence. Although cold stratification was not required for differentiation of the epicotyl–plumule, it was required to break the second part of PD, thereby allowing the shoot to emerge in spring.     

Angiotensin II induces complex fractionated electrogram in a cultured atrial myocyte monolayer mediated by calcium and sodium-calcium exchanger

Angiotensin II (AngII) has been implicated in the mechanism of atrial fibrillation (AF). There may be calcium-dependent pro-fibrillatory effect of AngII on atrial myocytes. We used cultured confluent HL-1 atrial myocyte monolayer with spontaneously propagated depolarization to study direct pro-fibrillatory effect of AngII and its molecular mechanism. AngII stimulation induced fibrillatory-like complex electrogram and calcium wave propagation. AngII shortened action potential duration and augmented calcium transient, thus increasing electrochemical gradient of forward-mode sodium-calcium exchanger (NCX) current and induced frequent irregular afterdepolarizations. AngII increased expression of sodium-calcium exchanger (NCX), further increasing calcium-membrane voltage coupling gain. The fibrillatory effect of AngII was attenuated by NCX blocker SEA0400 and NCX siRNA knockdown. AngII increased expression of L-type calcium channel and augmented calcium transient through PKC and CREB. The fibrillatory effect of AngII was also attenuated by PKC inhibitor chelerythrine and dominant negative form of CREB. In conclusions, AngII itself may electrically contribute to the mechanism of AF through increasing NCX expression and augmenting calcium transient, which is PKC and CREB dependent. Specific genetic knockdown of NCX attenuated calcium mediated afterdepolarization and complex electrogram.     

Binding kinetics of calmodulin with target peptides of three nitric oxide synthase isozymes

Efficient electron transfer from reductase domain to oxygenase domain in nitric oxide synthase (NOS) is dependent on the binding of calmodulin (CaM). Rate constants for the binding of CaM to NOS target peptides was only determined previously by surface plasmon resonance (SPR) (Biochemistry 35, 8742-8747, 1996) suggesting that the binding of CaM to NOSs is slow and does not support the fast electron transfer in NOSs measured in previous and this studies. To resolve this contradiction, the binding rates of holo Alexa 350 labeled T34C/T110W CaM (Alexa-CaM) to target peptides from three NOS isozymes were determined using fluorescence stopped-flow. All three target peptides exhibited fast k_on constants at 4.5 °C: 6.6 × 10⁸ M^− 1 s^− 1 for nNOS_726-749, 2.9 × 10⁸ M^− 1 s^− 1 for eNOS_492-511 and 6.1 × 10⁸ M^− 1 s^− 1 for iNOS_507-531, 3-4 orders of magnitude faster than those determined previously by SPR. Dissociation rates of NOS target peptides from Alexa-CaM/peptide complexes were measured by Ca²⁺ chelation with ETDA: 3.7 s^− 1 for nNOS_726-749, 4.5 s^− 1 for eNOS_492-511, and 0.063 s^− 1 for iNOS_507-531. Our data suggest that the binding of CaM to NOS is fast and kinetically competent for efficient electron transfer and is unlikely rate-limiting in NOS catalysis. Only iNOS_507-531 was able to bind apo Alexa-CaM, but in a very different conformation from its binding to holo Alexa-CaM.     

Micropropagation of bromeliad <Emphasis Type="Italic">Aechmea fasciata</Emphasis> via floral organ segments and effects of acclimatization on plantlet growth

Although suckers and seedlings can be used for the propagation of bromeliads, the low number of propagules and cross-variation limit their uniformity and mass cultivation. In this study, high-efficiency shoot organogenesis and plant regeneration were achieved on callus derived from petal and ovary explants of Aechmea fasciata (Bromeliaceae). Calluses were induced on half-strength Murashige and Skoog inorganic salts (1/2MS) supplemented with 1.0–1.5 mg l⁻¹ 2,4-dichlorophenoxyacetic acid in combination with 1.0 or 0.5 mg l⁻¹ α-naphthaleneacetic acid (NAA), and shoots regenerated after transfer to 1/2MS basal medium containing the combination of 1.0 mg l⁻¹ NAA + 0.5 mg l⁻¹ 1-phenyl-3-(1,2,3-thiadiazol-5-yl) urea. Those plantlets grown under a middle light intensity (50 μmol m⁻² s⁻¹) showed a dramatic increase in survival percentage (up to 95%) and the maximum number of newly developing roots. The plantlets that were transplanted onto pots were successfully grown in the greenhouse.     

Real-time PCR investigation on the expression of sboA and ituD genes in Bacillus spp

Aims: To investigate the expression of sboA and ituD genes among strains of Bacillus spp. at different pH and temperature. Methods and Results: Different Bacillus strains from the Amazon basin and Bacillus subtilis ATCC 19659 were investigated for the production of subtilosin A and iturin A by qRT‐PCR, analysing sboA and ituD gene expression under different culture conditions. Amazonian strains presented a general gene expression level lower than B. subtilis ATCC 19659 for sboA. In contrast, when analysing the expression of ituD gene, the strains from the Amazon, particularly P40 and P45B, exhibited higher levels of expression. Changes in pH (6 and 8) and temperature (37 and 42°C) caused a decrease in sboA expression, but increased ituD expression among strains from Amazonian environment. Conclusions: Temperature and pH have an important influence on the expression of genes sboA (subtilosin A) and ituD (iturin A) among Bacillus spp. The strains P40 and P45B can be useful for the production of antimicrobial peptide iturin A. Significance and Impact of the Study: Monitoring the expression of essential biosynthetic genes by qRT‐PCR is a valuable tool for optimization of the production of antimicrobial peptides.     

The effects of pad geometry and material properties on the biomechanical effectiveness of 26 commercially available hip protectors

Wearable hip protectors (padded garments) represent a promising strategy to decrease impact force and hip fracture risk during falls, and a wide range of products are currently marketed. However, little is known about how design features of hip protectors influence biomechanical effectiveness. We used a mechanical test system (simulating sideways falls) to measure the attenuation in femoral neck force provided by 26 commercially available hip protectors at three impact velocities (2, 3, and 4m/s). We also used a materials testing machine to characterize the force-deflection properties of each device. Regression analyses were performed to determine which geometric (e.g., height, width, thickness, volume) and force-deflection properties were associated with force attenuation. At an impact velocity of 3m/s, the force attenuation provided by the various hip protectors ranged between 2.5% and 40%. Hip protectors with lower stiffness (measured at 500N) provided greater force attenuation at all velocities. Protectors that absorbed more energy demonstrated greater force attenuation at the higher impact velocities (3 and 4m/s conditions), while protectors that did not directly contact (but instead bridged) the skin overlying the greater trochanter attenuated more force at velocities of 2 and 3m/s. At these lower velocities, the force attenuation provided by protectors that contacted the skin overlying the greater trochanter increased with increasing pad width, thickness, and energy dissipation. By providing a comparison of the protective value of a large range of existing hip protectors, these results can help to guide consumers and researchers in selecting hip protectors, and in interpreting the results of previous clinical trials. Furthermore, by determining geometric and material parameters that influence biomechanical performance, our results should assist manufacturers in designing devices that offer improved performance and clinical effectiveness.     

Honokiol inhibits LPS-induced maturation and inflammatory response of human monocyte-derived dendritic cells

Honokiol (HNK) is a phenolic compound isolated from the bark of houpu (Magnolia officinalis), a plant widely used in traditional Chinese and Japanese medicine. While substantial evidence indicates that HNK possesses anti-inflammatory activity, its effect on dendritic cells (DCs) during the inflammatory reaction remains unclear. The present study investigates how HNK affects lipopolysaccharide (LPS)-stimulated human monocyte-derived DCs. Our experimental results show that HNK inhibits the inflammatory response of LPS-induced DCs by (1) suppressing the expression of CD11c, CD40, CD80, CD83, CD86, and MHC-II on LPS-activated DCs, (2) reducing the production of TNF-α, IL-1β, IL-6, and IL-12p70 but increasing the production of IL-10 and TGF-β1 by LPS-activated DCs, (3) inhibiting the LPS-induced DC-elicited allogeneic T-cell proliferation, and (4) shifting the LPS-induced DC-driven Th1 response toward a Th2 response. Further, our results show that HNK inhibits the phosphorylation levels of ERK1/2, p38, JNK1/2, IKKα, and IκBα in LPS-activated DCs. Collectively, the findings show that the anti-inflammatory actions of HNK on LPS-induced DCs are associated with the NF-κB and mitogen-activated protein kinase (MAPK) signaling pathways.     

Characterization of TRPM8-like channels activated by the cooling agent icilin in the macrophage cell line RAW 264.7

Icilin is recognized as a chemical agonist of nociceptors and can activate TRPM8 channels. However, whether this agent has any effects on immune cells remains unknown. In this study, the effects of icilin on ion currents were investigated in RAW 264.7 murine macrophage-like cells. Icilin (1–100 μM) increased the amplitude of nonselective (NS) cation current (I _NS) in a concentration-dependent manner with an EC₅₀ value of 8.6 μM. LaCl₃ (100 μM) or capsazepine (30 μM) reversed icilin-induced I _NS; however, neither apamin (200 nM) nor iberiotoxin (200 nM) had any effects on it. In cell-attached configuration, when the electrode was filled with icilin (30 μM), a unique population of NS cation channels were activated with single-channel conductance of 158 pS. With the use of a long-lasting ramp pulse protocol, increasing icilin concentration produced a left shift in the activation curve of NS channels, with no change in the slope factor of the curve. The probability of channel opening enhanced by icilin was increased by either elevated extracellular Ca²⁺ or application of ionomycin (10 μM), while it was reduced by BAPTA-AM (10 μM). Icilin-stimulated activity is associated with an increase in mean open time and a reduction in mean closed time. Under current-clamp conditions, icilin caused membrane depolarization. Therefore, icilin interacts with the TRPM8-like channel to increase I _NS and depolarizes the membrane in these cells.