Does growth correlate with turgor-induced elastic strain in stems? A re-evaluation of de Vries' classical experiments

The correlation between growth and turgor-induced elastic expansion was studied in hypocotyls of sunflower (Helianthus annuus) seedlings under various growth conditions. Turgor-induced elastic cell wall strain was greater in hypocotyls of faster growing seedlings, i.e. in etiolated versus light-grown ones. It also was higher in rapidly growing young seedlings as compared with nongrowing mature ones. However, analysis of the spatial distribution of elastic strain and growth demonstrated that their correspondence was only apparent. Profiles of elastic strain declined steadily from the top of the hypocotyls toward the basis, whereas the profiles of relative elemental growth rate along the hypocotyls showed maxima within the growing zones. In contrast to earlier hypotheses, we conclude that turgor-induced elastic cell wall strain and growth do not correlate precisely in growing hypocotyls.     

Compensatory abilities depending on seasonal timing of thallus excision of the kelp Undaria pinnatifida cultivated in Matsushima Bay, northern Japan

Improved cultivation technology for the kelp Undaria pinnatifida is greatly needed to increase production to meet increasing commercial demand. A previous cultivation trial indicated that the crop yield of U. pinnatifida sporophytes could be increased greatly by thallus excision in late February due to compensatory growth of the remaining tissues. To develop this potential new cultivation technology, it is essential to identify the time period during which this kelp can compensate and its physiological responses to thallus excision. In this study, U. pinnatifida sporophytes were excised at about 30 cm above the meristem at the beginning of January, February, March, and April, respectively. Morphological features, photosynthesis, nutrient uptake, and carbon and nitrogen contents of excised kelps were measured and compared with these parameters in control kelps grown without excision. Both experimental and control kelps were farmed together in Matsushima Bay, northern Japan. The kelps excised in January and February showed significant increases in the lengths and dry weights of the blade, photosynthetic rates, nutrient uptake rates, and carbon and nitrogen contents compared with the control kelps, and the growth phase was prolonged for at least 1 month. No significant increases were found in dry weights and carbon and nitrogen contents of sporophylls until early April, which indicated that the maturation period was delayed. At the end of this experiment, the nitrogen contents of sporophyll tissues formed after excisions were significantly lower than those of tissues formed before excisions. In contrast, the kelps excised in March and April showed no significant increases in morphological and physiological parameters compared with control kelps. These results suggest that U. pinnatifida sporophytes exhibited great compensation when excisions were conducted during the growing phase in January and February but not in March and April when the maturation phase had started. The regulation of resource allocation to growth and maturation after thallus excisions in January and February likely results in prolongation of the growth phase and maturation phase in excised kelps.     

Compensation of the brown alga Undaria pinnatifida (Laminariales; Phaeophyta) after thallus excision under cultivation in Matsushima Bay, northern Japan

In recent years, the crop yield of cultivated Undaria pinnatifida (Harvey) Suringar has not been able to meet commercial demand and so advances in cultivation technology are strongly needed to increase production. Interestingly, cultivation work has shown the yield of U. pinnatifida sporophytes may be increased significantly by thallus excision, which may cause compensatory growth of the remaining tissues. To test this hypothesis and clarify correlative mechanisms, we examined seasonal morphological characteristics, photosynthetic rates, nutrient uptake rates, and carbon and nitrogen contents of U. pinnatifida sporophytes after thallus excision at a point 30 cm from the meristem in late February and compared these parameters with control kelps grown without excision, which were cultivated together in Matsushima Bay, northern Japan. Compared to control kelps, the length and dry weight of blades increased significantly after excision, and the growth phase was prolonged for about 1 month. The photosynthetic rates, nutrient uptake rates, and carbon and nitrogen contents of excised kelps were significantly higher than those of the controls. After vegetative growth stopped, resources accumulating in thalli were translocated significantly to sporophylls for maturation, indicating maturation was not negatively affected by thallus excision. These results indicate U. pinnatifida exhibits a very strong compensatory ability in response to thallus excision and consequently, the yield could be increased due to an increase in harvest frequency.     

Biochemical investigation of tissue growth: towards definition of "standard" tissue samples

This paper presents an animal model [the kangaroo], a quantitative anatomical dissection procedure, and a mathematical model [two-phase linear regression] which illustrate that body tissues grow at varying rates relative to each other. An argument is developed that biochemists interested in tissue chemical activity need to be able to sample tissue of known [predicted] growth rate. It is assumed that the ability to select, say muscle tissue samples, from any one animal at a stage of its growth where the individual selected pieces of tissue have known [predicted] low, average and high growth rates would allow comparisons to be made between the sampled tissues that may elucidate the underlying biochemical mechanisms involved in the growth process. It is asserted that to establish standards for tissue samples used in biochemical growth studies, the growth rate of the sampled tissue should be one of the criteria incorporated into the definition of what is "standard" for a tissue sample.     

Abscisic Acid accumulates at positive turgor potential in excised soybean seedling growing zones

Abscisic acid (ABA) accumulated in soybean (Glycine max [L.] Merr. cv Williams) hypocotyl elongating regions when seedlings were transferred to low water potential vermiculite (Ψ = −0.3 megapascals) even though positive turgor is retained in this tissue. Accumulation of ABA in growing zones could occur from de novo biosynthesis within this tissue or transport from adjacent nongrowing zones. Both growing and nongrowing hypocotyl and root tissues accumulated significant levels of ABA when excised and dehydrated to reduce turgor. Surprisingly, excised growing zones (which experienced no water loss) also accumulated ABA when incubated in darkness for 4 hours at 100% relative humidity and 29°C. Induction of ABA accumulation in the excised elongating region of the hypocotyl was not caused by disruption of root pressure or wounding. While excision of hypocotyl elongating regions induced ABA accumulation, no change in either extensin or p33 mRNA levels was observed. Accumulation of extensin or p33 mRNA required more severe wounding. This suggests that ABA is not involved in the response of these genes in wounded tissue and that wound signals are not causing ABA accumulation in excised tissue. Accumulation of ABA in excised elongating regions was correlated with growth inhibition and a decline in turgor to the yield threshold (Ψ;_p = 0.37 megapascals; R Matyssek, S Maruyama, JS Boyer [1988] Plant Physiol 86: 1163-1167). Inhibiting hypocotyl growth by transferring seedlings to lower temperatures or light did not cause ABA accumulation. We conclude that induction of ABA accumulation in growing zones is more sensitive to changes in turgor than the induction which occurs in mature tissues.     

Wound healing after excision of mantle tissue from the Akoya pearl oyster, Pinctada fucata

Pearl oysters are usually sacrificed to donate mantle tissue for pearl production. However, if oysters are anaesthetized, they are able to survive mantle excision and regenerate this tissue. Mantle excision causes a large wound and severs the pallial artery that necessitates rapid wound repair to avoid death by bleeding. This study was undertaken to assess the wound healing process in the mantle of the Akoya pearl oyster, Pinctada fucata, following mantle excision. Forty-seven P. fucata were relaxed with 2.5 mL L(-1) propylene phenoxetol before mantle tissue was excised. Oysters were relaxed and sacrificed 1, 3, 6, 12, 25, 36, 48, 66, 80 and 105 h after excision to assess mantle healing using histological techniques. Muscular contraction that effectively reduced the size of the wound was observed within 1 h after mantle excision. Accumulation of haemocytes and connective tissue occurred 3-6 h after excision and wound plugging was achieved within 6 h of excision. Proliferation of epithelial cells to cover the wound site was observed within the first 25 h after mantle excision and growth of connective tissue and formation of the pallial artery were observed within 105 h after mantle excision.     

Growth and betaine formation in Medicago sativa tissue cultures

Static and suspension tissue cultures of Medicago sativa were initiated from sterile seedlings, excised roots and aerial portions of the seedlings. Flask suspension cultures containing chlorophyll were also established. The various tissues were harvested after 30 days, growth rates were determined, and the tissue was later lyophilized. The lyophilized tissues from various batches were extracted separately with 70% EtOH. The betaine mixture was isolated from these extracts and from seeds of the plant as their tetraphenylborate salts. The betaines were separated from the latter complex via ion exchange and column chromatography and were identified by standard procedures. All of the tissues, regardless of origin, produced choline. Only chlorophyllous tissue biosynthesized stachydrine.     

同位素富集-稀释法研究食性转变对鱼类不同组织N同位素转化率的影响

稳定同位素技术广泛地用于描绘生态系统中食物网的食物来源和营养级关系,但是消费者不同组织转化率的研究相对较少。通过锦鲤摄食人工添加15N蓝藻的食性转化实验,研究不同组织N同位素转化率的差异,探讨组织生长和代谢对同位素转化的相对贡献,为不同时间尺度的稳定同位素研究取样奠定基础。结果表明,通过42d的加富蓝藻饲喂,各组织的N稳定同位素发生显著变化。肝的δ15N为(19.3±1.4)‰,显著高于其它组织,其次为鱼鳍((15.6±1.0)‰)和血液((12.6±0.4)‰),肌肉的δ15N‰最低,为(9.9±0.7)‰。在随后的同位素稀释实验中,锦鲤的体重增加,相对生长速率为0.011d-1,鳍肉的转化率最快,达到11.4%/d,半衰期仅为6.1d,其次是血液和肝,肌肉的转化率最低,仅有3.8%/d,半衰期最长,为18.4d。代谢衰减指数c和-1不存在显著差异,表明锦鲤各组织的N同位素转化主要由组织生长引起。结论显示,同位素富集-稀释法可以有效评价鱼类食性转变对不同组织同位素转化的差异,鳍肉和血液同位素分析可以作为锦鲤食性转变快速追踪的手段。     

A Test of Gas Exchange Measurements on Excised Canopy Branches of Ten Tropical Tree Species

We studied gas exchange of leaves on branches that had been cut and then re-cut under water to assess the utility of measuring gas exchange on leaves of excised canopy branches. There was large variation between species in their ability to photosynthesize following excision. Some species maintained up to 86.5% of intact photosynthetic rate 60 min after excision, whereas other species dropped below 40% of intact photosynthetic rates within 3 min. Three species showed significant reductions in maximum rates of gross photosynthetic rate (P _G) on leaves of excised branches relative to intact branches. Excision significantly reduced carboxylation rates (Vc _max) in four species and electron transport (J _max) in two species. There were also significant increases in compensation irradiance and reductions of day rates of respiration relative to intact measurements. While gas exchange on excised branches can provide useful measures for canopy species, responses of individual species to branch excision need to be taken into account. Measurements on pre-screened species allow a greater understanding of canopy photosynthesis of large trees when canopy access is not an option.     

Rapid wall relaxation in elongating tissues

Reported differences in the relaxation of cell walls in enlarging stem tissues of soybean (Glycine max [L.] Merr.) and pea (Pisum sativum L.) cause measurements of the yield threshold turgor, an important growth parameter, to be in doubt. Using the pressure probe and guillotine psychrometer, we investigated wall relaxation in these species by excising the elongating tissue in air to remove the water supply. We found that the rapid kinetics usually exhibited by soybean could be delayed and made similar to the slow kinetics previously reported for pea if slowly growing or mature tissue was left attached to the rapidly growing tissue when relaxation was initiated. The greater the amount of attached tissue, the slower the relaxation, suggesting that slowly growing tissue acted as a water source. Consistent with this concept was a lower water potential in the rapidly elongating tissue than in the slowly growing tissue. Previous reports of wall relaxation in pea included slowly growing tissue. If this tissue was removed from pea, relaxation became as rapid as usually exhibited by soybean. It is concluded that the true relaxation of cell walls to the yield threshold requires only a few minutes and that the yield threshold should be constant during so short a time, thus reflecting the yield threshold in the intact plant before excision. Under these conditions, the yield threshold was close to the turgor in the intact plant regardless of the species. The presence of slowly growing or mature tissue delays wall relaxation and should be avoided during such measurements. However, this delay can be used to advantage when turgor of intact growing tissues is being measured using excised tissues because turgor does not change for a considerable time after excision.     

Buckling of inner cell wall layers after manipulations to reduce tensile stress: observations and interpretations for stress transmission

The inner layer of the cell wall in tissues that are under tensile stress in situ, e.g. epidermis and collenchyma of etiolated sunflower hypocotyls, shows a pattern of transverse folds when the tissues are detached and plasmolysed. This can be observed by Nomarski imaging of inner surfaces of the outer cell walls and electron microscopy of longitudinal sections after peeling the epidermis and bathing it in plasmolysing solutions. The folds are apparently caused by buckling of the inner layer due to the longitudinal compressive force exerted on this layer by the outer wall layer, when it shrinks after the removal of the longitudinal tensile stresses. In these stresses, two components can be distinguished: the tissue stress, disappearing on peeling, and that caused directly by turgor pressure, disappearing in hyperosmotic solution. Investigation of the buckling indicates that the outer layer of the cell wall transmits in situ most of the longitudinal tensile stress in the wall. The common concept that the inner layer of the wall is the region bearing most stress and therefore regulating growth can still be valid with respect to the transverse stress component.     

Effects of Mechanical Stimulation on Avena Coleoptile Segment Elongation in a High Resolution, Continuous Growth-recording System

The effects of an abrasive mechanical stimulation of the inner epidermal surfaces of excised Avena coleoptile segments were examined in relation to growth in the presence and absence of exogenously supplied indole-3-acetic acid. Mechanical stimulation of this nature, provided immediately following excision, was found to elicit a small, transient increase in endogenous growth rate which contributed to a larger initial rapid growth response (previously referred to as a tactile response). These results, contrary to the earlier reports, suggest that the inner epidermal mechanical or tactile stimulation does not account for the entire initial rapid growth response. Preliminary experiments indicate that an alternative form of mechanical stimulation (segment excision) may contribute to that portion of initial rapid growth which is not attributable to inner epidermal abrasion.

Following its initial growth-enhancing effect, inner epidermal stimulation had either no effect or in some cases appeared inhibitory to endogenous growth. Growth in response to exogenous auxin was appreciably inhibited by this form of mechanical stimulation.

     

Growth response of testis tissue to partial castration in toads, Bufo bufo bufo

The growth response of the remaining intact testis or testis fragments to partial castration was studied as a function of the duration of the postoperative period, the amount of testis mass excised, as well as the functional state of the testes at the time of operation. Excision of about 90% of the testis mass caused a growth response that increased from slight after eight weeks to pronounced after 14 weeks. After 14 weeks the growth response was slight tounilateral excision of 75% of a testie and pronounced to bilateral excision of 75% of each testis. Subtotal castration caused formation of new seminiferous tubules within the remaining testis tissue when the operation was performed early in the annual testis cycle, whereas the growth response late in the cycle was primarily caused by increased spermatogenetic activity within existing tubules. Partial castration stimulated oocyte formation within fragmented testes, but not in a remaining intact testis. Oocyte formation within a testis fragment was independent of the presence of the Bidder's organ.     

Assessment of N-glycan heterogeneity of cactus glycoproteins by one-dimensional gel electrophoresis and matrix-assisted laser desorption/ionization time-of-flight mass spectrometry.

Artificial environmental conditions in tissue culture, such as elevated relative humidity and rich nutrient medium, can influence and modify tissue growth and induce spontaneous changes from characteristic organization pattern to unorganized callus. As succulent plants with crassulacean acid metabolism, cacti are particularly susceptible to this altered growth environment. Glycosylated proteins of Mammillaria gracillis tissues cultivated in vitro, separated by SDS-PAGE, were detected with Con A after the transfer of proteins onto the nitrocellulose membrane. The glycan components were further characterized by affinity blotting with different lectins (GNA, DSA, PNA, and RCA(120)). The results revealed significant differences in glycoprotein pattern among the investigated cactus tissues (shoot, callus, hyperhydric regenerant, and tumor). To test whether the N-glycosylation of the same protein can vary in different developmental stages of cactus tissue, the N-glycans were analyzed by MALDI-TOF MS after in-gel deglycosylation of the excised 38-kDa protein band. Paucimannosidic-type N-glycans were detected in oligosaccharide mixtures from shoot and callus, while the hyperhydric regenerant and tumor shared glycans of complex type. The hybrid oligosaccharide structures were found only in tumor tissue. These results indicate that the adaptation of plant cells to artificial environment in tissue culture is reflected in N-glycosylation, and structures of N-linked glycans vary with different developmental stages of Mammillaria gracillis tissues.     

Multimodality imaging in vivo for preclinical assessment of tumor-targeted doxorubicin nanoparticles

This study presents a new multimodal imaging approach that includes high-frequency ultrasound, fluorescence intensity, confocal, and spectral imaging to improve the preclinical evaluation of new therapeutics in vivo. Here we use this approach to assess in vivo the therapeutic efficacy of the novel chemotherapy construct, HerDox during and after treatment. HerDox is comprised of doxorubicin non-covalently assembled in a viral-like particle targeted to HER2+ tumor cells, causing tumor cell death at over 10-fold lower dose compared to the untargeted drug, while sparing the heart. Whereas our initial proof-of-principle studies on HerDox used tumor growth/shrinkage rates as a measure of therapeutic efficacy, here we show that multimodal imaging deployed during and after treatment can supplement traditional modes of tumor monitoring to further characterize the particle in tissues of treated mice. Specifically, we show here that tumor cell apoptosis elicited by HerDox can be monitored in vivo during treatment using high frequency ultrasound imaging, while in situ confocal imaging of excised tumors shows that HerDox indeed penetrated tumor tissue and can be detected at the subcellular level, including in the nucleus, via Dox fluorescence. In addition, ratiometric spectral imaging of the same tumor tissue enables quantitative discrimination of HerDox fluorescence from autofluorescence in situ. In contrast to standard approaches of preclinical assessment, this new method provides multiple/complementary information that may shorten the time required for initial evaluation of in vivo efficacy, thus potentially reducing the time and cost for translating new drug molecules into the clinic.     

Heat-induced changes in porcine annulus fibrosus biomechanics

The intervertebral disc is implicated as the source of low-back pain in a substantial number of patients. Because thermal therapy has been thought to have a therapeutic effect on collagenous tissues, this technique has recently been incorporated into several minimally invasive back pain treatments. However, patient selection criteria and precise definition of optimum dose are hindered by uncertainty of treatment mechanisms. The purpose of this study was to quantify acute changes in annulus fibrosus biomechanics after a range of thermal exposures, and to correlate these results with tissue denaturation. Intact annulus fibrosus (attached to adjacent vertebrae) from porcine lumbar spines was tested ex vivo. Biomechanical behavior, microstructure, peak of denaturation endotherm, and enthalpy of denaturation (mDSC) were determined before and after hydrothermal heat treatment at 37 degrees C, 50 degrees C, 60 degrees C, 65 degrees C, 70 degrees C, 75 degrees C, 80 degrees C, and 85 degrees C. Shrinkage of excised annular tissue (removed from adjacent vertebrae) was also measured after treatment at 85 degrees C. Significant differences in intact annulus biomechanics were observed after treatment, but the effects were much smaller in magnitude than those observed in excised annulus and those reported previously for other tissues. Consistent with this, intact tissue was only minimally denatured by treatment at 85 degrees C for 15 min, whereas excised tissue was completely denatured by this protocol. Our data suggest that in situ constraint imposed by the joint structure significantly retards annular thermal denaturation. These findings should aid the interpretation of clinical outcomes and provide a basis for the future design of optimum dosing regimens.     

Effect of plumule and radicle on somatic embryogenesis in the cultures of ginseng zygotic embryos

Immature zygotic embryos of ginseng produced somatic embryos on MS medium without growth regulators. However, in the culture of mature zygotic embryos, excision of the embryo was required for somatic embryo induction. Somatic embryos formed only on excised cotyledons without an embryo axis or on excised embryos without the plumule and radicle of the axis. This observation suggests that the axis tip of the embryo might suppress somatic embryo production although the cotyledon tissues have predetermined embryogenic competency. To clarify the role of the embryo axis on somatic embryo formation, excised plumules or radicles were placed in direct contact with the basal cut-ends of cotyledons. The adhesion of plumules or radicles highly suppressed somatic embryo formation from cotyledon explants. When an agar block containing exudate from excised plumules or radicles was placed in contact with the cut end of the cotyledon, a similar inhibition was observed. These results suggest that embryogenic competence is suppressed by endogenous inhibitors present in the axis tip of the zygotic embryo.     

The role of the apex in normal and tropic growth of sunflower hypocotyls

Regional growth in vertical and horizontal etiolated sunflower hypocotyls from which the apical hook tissue had been either partly or wholly excised, was measured 24 h later, the regions having been demarcated with resin beads. Removal of the cotyledons (an excision which included the distal end of the shoot apex) had little effect on growth during this period but excision of the apical hook significantly reduced growth. In vertically orientated seedlings, removal of half of the hook severely reduced growth in all other growing regions and removal of the entire hook totally inhibited growth. This inhibition of growth was not a consequence of the removal of the region of growth but a consequence of the removal of a region on which growth was dependent. In horizontal seedlings, the situation was more complex inasmuch as a horizontal orientation itself induced growth in previously non-growing regions. This new growth was localised in its extent and was not as severely affected by progressive excision of the hook as was growth in vertical seedlings. The results are discussed in terms of overall growth co-ordination in the hypocotyl.     

Stationarity of sodium channel gating kinetics in excised patches from neuroblastoma N1E 115.

Na channel gating parameters in a number of preparations are translated along the voltage axis in excised patches compared to cell attached or whole cell recording. The aim of this study is to determine whether these changes in gating behavior continue over an extended period or, rather, develop rapidly on excision with stationary kinetics thereafter. Average currents were constructed from single-channel records from neuroblastoma N1E 115 at various times after excision, excluding the first 5 min, in eight inside-out excised patches. Single exponentials were fitted to the current decay of the average records, and the mean time constant for each patch was determined. Values were plotted as the percentage difference from these means for each patch against time from excision. Collected results show no obvious trend in values from 5 min to 2 h. Kinetics are stationary, and shifts in Na channel gating parameters along the voltage axis seen in excised as compared to whole cell configuration in neuroblastoma must be complete by the first few minutes after excision. Raising the internal Na concentration reduced the single channel current amplitude, confirming that these are Na channels.     

Accumulation of phosphate, sulfate and sucrose by excised Phloem tissues

Excised petiolar vascular bundles and excised phloem tissues have been shown to take up phosphate, sulfate and sucrose by a true accumulation process and against high concentration ratios. Phosphate was accumulated principally as inorganic phosphate, and sucrose principally as sucrose. The rates of accumulation of the 3 solutes into the phloem-containing tissues were from 4 to 35 times higher than into comparable parenchyma tissue. It is suggested that this active accumulation mechanism plays an important role in the phenomenon of phloem transport.

The excised vascular, phloem and parenchyma tissues show an aging phenomenon: aerating the excised tissues for 18 hours prior to their use causes marked changes in the accumulatory behavior of the tissue. The data suggest that 1 phosphate accumulation system of low affinity but high capacity exists in fresh tissue, and that aging allows the development of a second, additional phosphate accumulation mechanism of high affinity and low capacity. A possible role in the control of phosphate movement is suggested.