The role of the epidermis as a stiffening agent in Tulipa (Liliaceae) stems

We investigated the hypothesis that the epidermis is a tension-stressed "skin' whose contribution to stem stiffness depends on the turgor pressure exerted on it by an hydrostatically inflated inner "core' of tissues. This hypothesis was tested by relying on the intensities of bending stresses due to stem flexure, which must reach their maximum levels at the outer surface of epidermis such that damage to the surface of the stem should produce the most significant decrease in overall flexural stiffness. We discerned whether the principal tension supporting members at the stem surface (cellulosic microfibrils) were oriented parallel or normal to stem length by comparing the bending stiffness of stems before and after their surface cells first received three parallel longitudinal incisions followed by one helical incision, and by comparing the bending stiffness of stems for which the sequence of cuts was reversed. The same protocol was also applied to stems with various water potentials to determine the effect of hydrostatic pressure on stem stiffness contributed by the surface. Based on the behavior of 82 turgid Tulipa stems, parallel cuts reduced, on average, stem stiffness by 8%, whereas a subsequent helical incision further reduced stiffness by 42%. In contrast, an initial helical incision reduced stem stiffness by 50%, while three subsequent parallel cuts through the same stems did not significantly further reduce stiffness. These results suggested that the net orientation of cellulose microfibrils in the outer epidermal walls was parallel to stem length. This was confirmed by microscopic observations of cells with dichroic staining and polarized light. The responses to surgical damage were directly proportional to stem water potential. We thus conclude that the epidermis, probably in conjunction with a single layer of subepidermal collenchyma cells, acts as a tension-stiffening agent that can contribute as much as 50% to overall stem stiffness We present a simple mechanical model that can account for all our observations.     

Axis elongation can occur with net longitudinal orientation of wall microfibrils

During the initial phases of elongation of pea internodes, oat and rice coleoptiles, oat mesocotyls, soybean hypocotyls and dandelion peduncles, net transverse orientation of cellulose wall microfibrils (Mfs) was found in the outer epidermal wall. This paper demonstrates that in all these axes, with the exception of rice coleoptile, net longitudinal orientation of microfibrils occurs in the outer epidermal wall in portions of the axes that were still elongating at the time of sampling. The timing of the transition to net longitudinal orientation and whether the transition proceeded acropetally or basipetally varied with the type of axis under study. The variability of the relationship between extension and the transition from net transverse to net longitudinal orientation suggests that factors other than extension are important in determining the transition. Layers of longitudinal wall microfibrils may be added to the extending epidermal wall to bolster its tensile strength commensurate with its function during and after extension. Attention is drawn to the parallels between the concept of tissue tension in growing axes and the concept that the epidermis functions as a stressed skin in the support of mature plant parts in primary growth.     

Biomechanics of the columnar cactus Pachycereus pringlei

We report the longitudinal variations in stiffness and bulk density of tissue samples drawn from along the length of two Pachycereus pringlei plants measuring 3.69 and 5.9 m in height to determine how different tissues contribute to the mechanical stability of these massive vertical organs. Each of the two stems was cut into segments of uniform length and subsequently dissected to obtain and mechanically test portions of xylem strands, stem ribs, and a limited number of pith and cortex samples. In each case, morphometric measurements were taken to determine the geometric contribution each tissue likely made to the ability of whole stems to resist bending forces. The stiffness of each xylem strand increased basipetally toward the base of each plant where stiffness sharply decreased, reaching a magnitude comparable to that of strands 1 m beneath the stem apex. The xylem was anisotropic in behavior, i.e., its stiffness measured in the radial and in the tangential directions differed significantly. Despite the abrupt decrease in xylem strand stiffness at the stem base, the contribution made by this tissue to resist bending forces increased exponentially from the tip to the base of each plant due to the accumulation of wood. A basipetal increase in the stiffness of the pith (and, to limited extent, that of the cortex) was also observed. In contrast, the stiffness of stem rib tissues varied little as a function of stem length. These tissues were stiffer than the xylem in the corresponding portions of the stem along the upper two-fifths of the length of either plant. Tissue stiffness and bulk density were not significantly correlated within or across tissue types. However, a weak inverse relationship was observed for these properties in the case of the xylem and stem rib tissues. We present a simple formula that predicts when stem ribs rather than the xylem strands serve as the principal stiffening agents in stems. This formula successfully predicted the observed aspect ratio of the stem ribs (the average quotient of the radial and tangential dimensions of rib transections), and thus provided circumstantial evidence that the ribs are important for mechanical stability for the distal and younger regions of the stems examined.     

Tissue stresses in growing plant organs

Rapidly growing plant organs (e.g. coleopties, hypocotyls, or internodes) are composed of tissues that differ with respect to the thickness, structure, and extensibility of their cell walls. The thick, relatively inextensible outer wall of the epidermal cells contains both transverse and longitudinally oriented cellulose-microfibrils. The orientation of microfibrils of the thin, extensible walls of the parenchyma cells seems to be predominantly transverse. In many growing organs (i.e. leafstalks), the outer epidermal wall is supported by a thickened inner epidermal wall and by thick-walled subepidermal collenchyma tissue. Owing to the turgor pressure of the cells the peripheral walls are under tension, while the extensible inner tissue is under compression. As a corollary, the longitudinal tensile stress of the rigid peripheral wall is high whereas that of the internal walls is lowered. The physical stress between the tissues has been described by Sachs in 1865 as 'tissue tension'. The term 'tissue stress'. however, seems to be more appropriate since it comprises both tension and compression. Hitherto no method has been developed to measure tissue stresses directly as force per unit cross-sectional area. One can demonstrate the existence of tissue stresses by separation of the tissues (splitting, peeling) and determining the resulting strain of the isolated organ fragments. Based on such experiments it has been shown that rapid growth is always accompanied by the existence of longitudinal tissue stresses.     

The Role of the Epidermis in the Control of Elongation Growth in Stems and Coleoptiles

The peripheral cell wall(s) of stems and coleoptiles are 6 to 20 times thicker than the walls of the inner tissues. In coleoptiles, the outer wall of the outer epidermis shows a multilayered, helicoidal cellulose architecture, whereas the walls of the parenchyma and the outer wall of the inner epidermis are unilayered. In hypocotyls and epicotyls both the epidermal and some subepidermal walls are multilayered, helicoidal structures. The walls of the internal tissues (inner cortex, pith) are unilayered, with cellulose microfibrils oriented primarily transversely. Peeled inner tissues rapidly extend in water, whereas the outer cell layer(s) contract on isolation. This indicates that the peripheral walls limit elongation of the intact organ. Experiments with the pressure microprobe indicate that the entire organ can be viewed as a giant, turgid cell: the extensible inner tissues exert a pressure (turgor) on the peripheral wall(s), which bear the longitudinal wall stress of the epidermal and internal cells. Numerous studies have shown that auxin induces elongation of isolated, intact sections by loosening of the growth-limiting peripheral cell wall(s). Likewise, the effect of light on reduction of stem elongation and cell wall extensibility in etiolated seedlings is restricted to the peripheral cell layers of the organ. The extensible inner tissues provide the driving force (turgor pressure), whereas the rigid peripheral wall(s) limit, and hence control, the rate of organ elongation.     

Changes in microfibril arrangement on the inner surface of the epidermal cell walls in the epicotyl of Vigna angularis ohwi et ohashi during cell growth

Throughout the entire period of cell growth, the microfibrils on the inner surface of the outer tangential walls of the epidermal cells of Vigna angularis epicotyls are running parallel to one another and their orientation differs from cell to cell. Although transverse, oblique and longitudinal microfibrils can be observed irrespective of cell age, the frequency distribution of microfibril orientation changes with age. In young cells, transversely oriented microfibrils predominate. In cells of medium age, which are still undergoing elongation, transverse, oblique and longitudinal microfibrils are present in quite similar frequencies. In old, non-growing cells, longitudinally oriented microfibrils are predominent. A decrease in the relative frequency of transversely oriented microfibrils with cell age was also observed in the radial epidermal walls.     

Comparisons among biomass allocation and spatial distribution patterns of some vine,pteridophyte, and gymnosperm shoots

The interspecific allometry of leaf, stem, and reproductive biomass distal to stem diameter was determined for a total of 12 angiosperm vine, gymnosperm, and pteridophyte species to compare allocation patterns to vegetative and reproductive shoot organs. The allometry of stem diameter in terms of the distance from shoot apices also was determined to quantify the manner in which vines, gymnosperms, and pteridophyte stems tapered along their length. The stems of vine species were found to weigh more than those of arborescent gymnosperm species distal to any point of equivalent stem diameter. Vine species also distribute more of their stem mass to shoot length as opposed to girth than gymnosperm species. Vine stems also supported proportionally larger leaf and reproductive biomass in comparison to gymnosperm stems of equivalent diameter, yet partitioned their total shoot biomass more or less equally between leaf and stem biomass in the same manner as the gymnosperm species examined. The allometry of vine as well as gymnosperm leaf biomass with respect to stem biomass appeared to be slightly anisometric and negative, suggesting that more massive stems had proportionally less leaf biomass than their smaller, less massive counterparts. Vine stems could be approximated as very slender cones; the shape and geometry of gymnosperm stems complied with those of stubby, truncated cones whose top diameter (for those examined), on the average, equaled 28% of the basal diameter. In general terms, the interspecific allometry of vines was most similar to that of pteridophytes. Collectively, these data refute the commonly held notion that vine stems are simply more slender than those of species with self-supporting stems.     

Biomechanics and functional anatomy of hollow-stemmed sphenopsids. I. Equisetum giganteum (Equisetaceae)

The hollow stem of Equisetum giganteum owes its mechanical stability to an outer ring of strengthening tissue, which provides stiffness and strength in the longitudinal direction, but also to an inner lining of turgid parenchyma, which lends resistance to local buckling. With a height >2.5 m isolated stems are mechanically unstable. However, in dense stands individual stems support each other by interlacing with their side branches, the typical growth habit of semi-self-supporters.     

Arrangement of Cellulose Microfibrils in Walls of Elongating Parenchyma Cells

The arrangement of cellulose microfibrils in walls of elongating parenchyma cells of Avena coleoptiles, onion roots, and celery petioles was studied in polarizing and electron microscopes by examining whole cell walls and sections. Walls of these cells consist firstly of regions containing the primary pit fields and composed of microfibrils oriented predominantly transversely. The transverse microfibrils show a progressive disorientation from the inside to the outside of the wall which is consistent with the multinet model of wall growth. Between the pit-field regions and running the length of the cells are ribs composed of longitudinally oriented microfibrils. Two types of rib have been found at all stages of cell elongation. In some regions, the wall appears to consist entirely of longitudinal microfibrils so that the rib forms an integral part of the wall. At the edges of such ribs the microfibrils can be seen to change direction from longitudinal in the rib to transverse in the pit-field region. Often, however, the rib appears to consist of an extra separate layer of longitudinal microfibrils outside a continuous wall of transverse microfibrils. These ribs are quite distinct from secondary wall, which consists of longitudinal microfibrils deposited within the primary wall after elongation has ceased. It is evident that the arrangement of cellulose microfibrils in a primary wall can be complex and is probably an expression of specific cellular differentiation.     

Non-parametric statistical formulas for factors of safety of plant stems

A previously proposed statistical approach for computing factors of safety (i.e. numerical measures of mechanical reliability) for any load bearing structure, like a vertical plant stem, is here extended to cope with organic structures whose morphological or mechanical properties have Weibull frequency distributions. This approach is illustrated using the actual length L and critical buckling length Lerof flower stalks (peduncles) collected from isogenic garlic (Allium sativum) populations grown under windy field and protected glasshouse conditions. Our analyses of the data indicate that L and Lerof peduncles harvested from both populations have Weibull frequency distributions, that the factor of safety for glasshouse grown peduncles is very near unity (i.e. S=1.03), and that the factor of safety of field grown peduncles is 73% higher than that of glasshouse grown plants (i.e. S=1.73). Comparisons between the S -values computed on the basis of our formulas and on the basis of the quotient of the mean values of Lerand L for each of the two populations indicate that the statistical method gives biologically realistic S -values and that the difference in the S -values for stems grown under protected and unprotected environmental conditions likely reflects the effects of chronic mechanical perturbation (due to wind-induced drag) on normal stem growth and development. Copyright 1999 Academic Press.     

Biomechanics and Functional Anatomy of Hollow Stemmed Sphenopsids: III. Equisetum hyemale

As in Equisetum giganteum, the hollow stem of Equisetum hyemale owes the mechanical stability of the internodes to an outer ring of strengthening tissue (hypodermal sterome) which provides stiffness and strength in the longitudinal direction. In contrast to hollow-stemmed grasses, the hypodermal sterome consists of living cells. The compound inner tissue of the overwintering aerial stem of Equisetum hyemale includes a continuous inner and outer endodermis layer of vital thick-walled cells that have slightly lignified Casparian thickenings. The two endodermis layers provide an inner tension and compression bracing which lend resistance to local buckling. The stress-strain relation in longitudinal tension is biphasic with remarkably high critical strains especially in the upper parts of the stem. Scraping off part of the epidermis with the built-in silicate does not change this behaviour, except in the initial steep part of the curve where the Young's modulus is reduced by 20%. No contribution of the endodermis and the parenchyma could be detected in tension tests of longitudinally-oriented strips. Relaxation experiments reveal viscoelastic behaviour. As with the biphasic stress-strain relation and the critical strains, the viscoelastic properties have largely to be ascribed to the hypodermal sterome.     

SOME OBSERVATIONS ON THE MECHANISM OF ORIENTATION MOVEMENT OF WOODY STEMS

The force that induces orientation movement of inclined or bent woody stems is generated in reaction wood even in young terminal stems with a large proportion of soft tissues to secondary xylem. Compression wood formed in pine as a response to inclination expands longitudinally after the stresses are released by sawing it from the stem. The increment of length of compression wood when sawed is equal to the decrement of its length which occurs during drying. This suggests that stresses developed by compression wood in the stem are related to imbibition of water by its cell walls. Not all compression wood develops tensile forces in the stem. Neutral compression wood was observed in the lower portion of inclined stems of pine. Tension wood in poplar develops contractile forces in the stem during its aestival maturation. However, when harvested before developing contractile forces in situ, it develops such forces during drying. This suggests that in poplar the mechanism which produces forces responsible for orientation bending also involves changes in cell wall hydration.     

Trubs,but no trianas: filled and empty regions of angiosperm stem length‐diameter‐mechanics space

Discrete plant habit categories such as ‘tree’, ‘shrub’, and ‘liana’ belie continuous variation in nature. To study the evolution of this continuous variation, we gathered data on stem length, diameter and tissue mechanical stiffness across a highly morphologically diverse highland xerophytic scrub on a lava flow in central Mexico. With stem allometric and mechanical data from 1216 segments from 50 species, we examined relationships between stem length–diameter proportions and tissue mechanical stiffness using linear mixed‐effects models. Rather than a series of discrete clouds in stem length–diameter–tissue stiffness space, corresponding to traditional habit categories, the plants of this xerophytic scrub formed a single continuous one. Within this cloud, self‐supporting plants had stems that became predictably longer and tissues that became stiffer for a given diameter increase, and there was no paucity of intermediates between trees and shrubs (‘trubs’). Non self‐supporting plants had a steeper stem length–diameter slope and their tissues did not increase in stiffness with stem size. The area between self‐ and non self‐supporting plants was sparsely occupied as stem size increased. We predict that this ‘empty’ space between lianas and trees is developmentally accessible but of low fitness, meaning that there should be few ‘trianas’ in nature. © 2015 The Linnean Society of London, Botanical Journal of the Linnean Society, 2015, 179 , 361–373.     

Effects of tension and transmural stimulation on prostaglandin production by estrous rabbit oviductal isthmus

The effect of longitudinal and circular stretch on the amounts of Prostaglandin F (PGF) and Prostaglandin E (PGE) found in the fluid bathing rabbit oviductal isthmus has been investigated. It was found that the amounts of PGE nad PGF measured in the bathing fluid of longitudinally or circularly stretched tissues were negatively correlated to the maturity of the animal. Prostaglandin E increased with time in the tissues under longitudinal and circular tension. Prostaglandin F also increased with time under longitudinal tension but remained fairly constant under circular tension. Increasing the load from 0.5 to 2.0 g had no significant effect on PGE found under longitudinal or circular tension or on PGF found under longitudinal tension. Under circular tension, PGF found increased. Transmural stimulation at 20 Hz increased PGE 8-fold over control values while PGF increased only 1 to 3-fold. It is suggested that distension of the rabbit oviductal isthmus results in increased PGF production, which could be important in ovum transport.     

THE STRUCTURE OF THE PRIMARY EPIDERMAL CELL WALL OF AVENA COLEOPTILES

The primary walls of epidermal cells in Avena coleoptiles ranging in length from 2 to 40 mm. have been studied in the electron and polarizing microscopes and by the low-angle scattering of x-rays. The outer walls of these cells are composed of multiple layers of cellulose microfibrils oriented longitudinally; initially the number of layers is between 10 and 15 but this increases to about 25 in older tissue. Where epidermal cells touch, these multiple layers fuse gradually into a primary wall of the normal type between cells. In these radial walls, the microfibrils are oriented transversely. Possible mechanisms for the growth of the multilayered outer wall during cell elongation are discussed.     

Light-induced inhibition of elongation growth in sunflower hypocotyls

Summary The long-term effects of white light (WL) on epidermal cell elongation and the mechanical properties and ultrastructure of cell walls were investigated in the subapical regions of hypocotyls of sunflower seedlings (Helianthus annuus L.) that were grown in darkness. Upon transition to WL a drastic inhibition of epidermal cell elongation was observed. However, the mechanical properties of the inner tissues (cortex, vascular bundles, and pith) were unaffected by WL. Thus, the light-induced decrease in cell wall plasticity measured on entire stems occurs exclusively in the peripheral tissues (epidermis and 2 to 3 subepidermal cell layers).An electronmicroscopic investigation of the epidermal cell walls showed that they are of the helicoidal type with the direction of microfibrils monotonously changing during deposition. This cell wall type was identified by the appearance of arced patterns of microfibrils in cell walls sectioned oblique to the plane of their synthesis. WL irradiation did not change the periodicity of this pattern nor the thickness of the lamellae. Thus, the inhibition of cell elongation was not caused or accompanied by a shift in the direction of microfibril deposition in the growth-limiting outer tissues. However, cell wall thickness, the number of lamellae and hence the amount of cellulose oriented parallel and transverse to the longitudinal cell axis increased in WL. This may account for the effect of WL on the reduction of cell wall plasticity and growth.Abbreviations D darkness - PATAg periodic acid-thiocarbohydracide-silver protein - WL white light     

Cell growth pattern and wall microfibrillar arrangement: experiments with nitella

In cylindrical cells growing throughout their length, over-all transverse reinforcement of the wall by microfibrils is believed to be required for cell elongation. The multinet theory states that in such cells microfibrils are deposited at the inner surface of the wall with transverse orientation and are then passively reoriented toward the longitudinal direction by the predominant longitudinal strain (surface expension). In the present study young Nitella cells were physically forced to grow in highly abnormal patterns: in length only, in girth only, or with localized suppression of growth. Subsequent gradients of microfibrillar arrangement within the wall cross-section were measured with polarized light and interference microscopes. The novel wall structures produced were in all cases explainable by passive reorientation, i.e. by the multinet theory. The study also showed that orientation of synthesis remains insensitive to several of the physical manipulations that strongly influence the passive behavior of wall microfibrils. Only the localized complete suppression of surface growth led to the deposition of nontransverse cellulose. These results suggest that the presence of strain is needed for continued oriented synthesis, but that the directional aspect of strain is not an “instructional” agent continuously guiding the orientation of synthesis, once this orientation has been established.     

Ontogenetic tissue modification in Malus fruit peduncles: the role of sclereids

Background and Aims

Apple (Malus) fruit peduncles are highly modified stems with limited secondary growth because fruit ripening lasts only one season. They must reliably connect rather heavy fruits to the branch and cope with increasing fruit weight, which induces dynamic stresses under oscillating wind loads. This study focuses on tissue modification of these small, exposed structures during fruit development.

Methods

A combination of microscopic, static and dynamic mechanical tests, as well as Raman spectroscopy, was used to study structure–function relationships in peduncles of one cultivar and 12 wild species, representatively chosen from all sections of the genus Malus. Tissue differentiation and ontogenetic changes in mechanical properties of Malus peduncles were observed throughout one growing season and after successive removal of tissues.

Key Results

Unlike in regular stems, the vascular cambium produces mainly phloem during secondary growth. Hence, in addition to a reduced xylem, all species developed a centrally arranged sclerenchyma ring composed of fibres and brachysclereids. Based on differences in cell-wall thickness, and proportions and arrangement of sclereids, two types of peduncle construction could be distinguished. Fibres provide an increased maximum tensile strength and contribute most to the overall axial rigidity of the peduncles. Sclereids contribute insignificantly to peduncle strength; however, despite being shown to have a lower elastic modulus than fibres, they are the most effective tissue in stiffening peduncles against bending.

Conclusions

The experimental data revealed that sclereids originating from cortical parenchyma act as ‘accessory’ cells to enhance proportions of sclerenchyma during secondary growth in peduncles. The mechanism can be interpreted as an adaptation to continuously increasing fruit loads. Under oscillating longitudinal stresses, sclereids may be regarded as regulating elements between maintenance of stiffness and viscous damping, the latter property being attributed to the cortical parenchyma.     

Gibberellin-induced formation of tension wood in angiosperm trees

After gibberellin had been applied to the vertical stems of four species of angiosperm trees for approximately 2 months, we observed eccentric radial growth that was due to the enhanced growth rings on the sides of stems to which gibberellin had been applied. Moreover, the application of gibberellin resulted in the formation of wood fibers in which the thickness of inner layers of cell walls was enhanced. These thickened inner layers of cell walls were unlignified or only slightly lignified. In addition, cellulose microfibrils on the innermost surface of these thickened inner layers of cell walls were oriented parallel or nearly parallel to the longitudinal axis of the fibers. Such thickened inner layers of cell walls had features similar to those of gelatinous layers in the wood fibers of tension wood, which are referred to as gelatinous fibers. Our anatomical and histochemical investigations indicate that the application of gibberellin can induce the formation of tension wood on vertical stems of angiosperm trees in the absence of gravitational stimulus.