Anatomical and ultrastructural examination of adventitious root formation in stem slices of apple

Adventitious root formation in vitro in 1-mm stem slices cut from microshoots of apple cv. Jork 9 was studied using light and electron microscopy. When indole-3-butyric acid (IBA) had been added to the medium, starch grains accumulated during the first 24 h of culture in cells of the cambial region and in cells in the vicinity of vascular tissue and in the primary rays. This accumulation occurred only in the basal part of explants. After that, the nuclei in these cells were activated, and the density of the cytoplasm and the number of cell organelles increased, whereas starch was broken down. Cambium cells started to divide transversely and at 96 h, after several divisions, a continuous ring of isodiametric cytoplasmic cells had appeared around the xylem near the basal cutting surface. The cells in this ring were rich in cell structures, and did not contain large starch grains and a central vacuole. Root meristemoids regenerated from the portions of the ring that were localized in the primary rays. From the other cells in the ring, callus developed. The meristemoids did not grow into the direction of the epidermis as in shoots, but along the vascular bundles. After emergence from the cutting surface, the meristemoids were transformed into small, dome-like primordia. They developed a typical root apex with root cap, root ground meristem and tracheid connection with shoot vascular tissue. This revised version was published online in July 2006 with corrections to the Cover Date.     

The monoclonal antibody 22/18 recognizes a conformational change in an intermediate filament of the newt,Notophthalmus viridescens,during limb regeneration

Summary Previous work has shown that the monoclonal antibody 22/18 identifies progenitor cells (blastemal cells) which depend on the nerve for their division in the early stages of limb regeneration in the newt,Notophthalmus viridescens. This antibody also reacts with cultured cells derived from the newt limb, and the intensity of immunoreactivity appears related to cell density and differentiation into myotubes. We report here that the monoclonal antibody 22/18 recognizes a polypeptide (22/18 antigen) which is intracellular and filamentous. Double staining of cells with 22/18 monoclonal antibody and antibodies against various cytoskeletal components indicates that the epitope is expressed on an intermediate filament component. Although this antibody is specific for blastemal cells in cryostat sections of the regenerating limb, its reactivity on immunoblots is not confined to this tissue. The 22/18 antigen is differentially affected by aldehyde fixatives distinguished by the spacing of their reactive groups. While formaldehyde fixation impairs detection of the antigen, ethylene glycol-bis[succinic acid n-hydroxysuccinimide ester] reveals the antigen in sections of normal and regenerating limbs in a distribution that is consistent with the one obtained from immunoblots. We suggest that the 22/18 monoclonal antibody detects a change in protein conformation, probably related to changes in the physiological state of the cell, that occurs transiently during regeneration and possibly during development.     

Dual role of osteoblastic proenkephalin derived peptides in skeletal tissues

Proenkephalin encodes a group of small peptides with opiate-like activity, the endogenous opioids, known to function as neurohormones, neuromodulators, and neurotransmitters. Recently, we have demonstrated that in addition to its abundance in fetal brain tissue, proenkephalin is highly expressed in nondifferentiated mesodermal cells of developing fetuses. We identified the skeletal tissues, bone, and cartilage as major sites of proenkephalin expression. To examine the possibility that proenkephalin is involved in bone development we have studied the expression of this gene in bone-derived cells, its modulation by bone active hormones, and the effects of enkephalin-derived peptides on osteoblastic phenotype. Our studies revealed that osteoblastic cells synthesize high levels of proenkephalin mRNA which are translated, and the derived peptides are secreted. Reciprocal interrelationships between osteoblast maturation and proenkephalin expression were established. These results together with our observations demonstrating inhibitory effects of proenkephalin-derived peptides on osteoblastic alkaline phosphatase activity, strongly support the notion that proenkephalin is involved in bone development. A different direction of research by other investigators has established the capability of the opioid system in the periphery to participate in the control of pain. On the basis of these two lines of observation, we would like to present the following hypothesis: The potential of embryonic skeletal tissue to synthesize proenkephalin-derived peptides is retained in the adult in small defined undifferentiated cell populations. This potential is realized in certain situations requiring rapid growth, such as remodeling or fracture repair. We suggest that in these processes, similarly to the situation in the embryo, the undifferentiated dividing cells produce the endogenous opioids. In the adult these peptides may have a dual function, namely participating in the control of tissue regeneration and in the control of pain. © 1994 Wiley-Liss, Inc.     

Wound phloem in transition to bundle phloem in primary roots ofPisum sativum L.

Summary 48 hours after interrupting the root stele ofPisum, wound phloem initiated (proximally or distally to the wound) to reconnect the vascular stumps was found to contain some nucleate wound-sieve elements. At the elongating end of an incomplete wound-sieve tube these elements exhibit a sequence of ultrastructural changes as known from protophloem-sieve tubes. Elongation occurs by the addition of newly divided (wound-) sieve-element/companion-cell complexes. In order to dedifferentiate and assume a new specialization formerly quiescent stelar or cortical cells require at least one (mostly more) preliminary division. Companion cells are consequently obligatory sister cells to wound-sieve elements.By reconstruction using serial sections it could be shown that wound-sieve tubes elongate bidirectionally, starting in an early activated procambial cell of the stele. The elongation is directed by the existence of plasmodesmata, preferably when lying in primary pit fields, and by the plane of preceding divisions. Thus, the developing wound-sieve tube can deviate from the damaged bundle and radiate into the cortex as soon as the plane of the preceding divisions is favourable. In the opposite direction, elongating wound-sieve tubes run parallel to pre-existing phloem traces, thus broading their base at the bundle for the deviating part of the wound-sieve tube. Frequently an individual wound-sieve tube is supplemented at the bundle by a further wound-sieve tube which is partly running parallel to it. Both sieve tubes are interlinked with sieve plates by three-poled sieve elements.Ultrastructurally, the developmental changes of nucleate wound-sieve elements follow the known pattern. In spite of its contrasting origin and odd shape a mature wound-sieve element eventually has the same contents as regular sieve elements: sieve-element plastids, mitochondria, stacked ER and small amounts of P-protein within an electronlucent cytoplasm.     

Cardiac Restricted Overexpression of Membrane Type-1 Matrix Metalloproteinase Causes Adverse Myocardial Remodeling following Myocardial Infarction

The membrane type-1 matrix metalloproteinase (MT1-MMP) is a unique member of the MMP family, but induction patterns and consequences of MT1-MMP overexpression (MT1-MMPexp), in a left ventricular (LV) remodeling process such as myocardial infarction (MI), have not been explored. MT1-MMP promoter activity (murine luciferase reporter) increased 20-fold at 3 days and 50-fold at 14 days post-MI. MI was then induced in mice with cardiac restricted MT1-MMPexp (n = 58) and wild type (WT, n = 60). Post-MI survival was reduced (67% versus 46%, p < 0.05), and LV ejection fraction was lower in the post-MI MT1-MMPexp mice compared with WT (41 ± 2 versus 32 ± 2%,p < 0.05). In the post-MI MT1-MMPexp mice, LV myocardial MMP activity, as assessed by radiotracer uptake, and MT1-MMP-specific proteolytic activity using a specific fluorogenic assay were both increased by 2-fold. LV collagen content was increased by nearly 2-fold in the post-MI MT1-MMPexp compared with WT. Using a validated fluorogenic construct, it was discovered that MT1-MMP proteolytically processed the pro-fibrotic molecule, latency-associated transforming growth factor-1 binding protein (LTBP-1), and MT1-MMP-specific LTBP-1 proteolytic activity was increased by 4-fold in the post-MI MT1-MMPexp group. Early and persistent MT1-MMP promoter activity occurred post-MI, and increased myocardial MT1-MMP levels resulted in poor survival, worsening of LV function, and significant fibrosis. A molecular mechanism for the adverse LV matrix remodeling with MT1-MMP induction is increased processing of pro-fibrotic signaling molecules. Thus, a proteolytically diverse portfolio exists for MT1-MMP within the myocardium and likely plays a mechanistic role in adverse LV remodeling.     

Does Weeding Promote Regeneration of an Indigenous Tree Community in Felled Pine Plantations in Uganda?

The use of plantations to manage extensive tracks of deforested lands in the tropics is a conservation strategy that has recently received considerable attention. Plantation trees can promote seed dispersal by attracting dispersers and creating favorable site conditions, leading to increased germination and establishment of indigenous trees. Subsequently, plantation trees can be harvested for profit or left to senesce, leaving a native tree community. We evaluated the effect of vine, grass, and shrub cutting (weeding) over a 3‐year period on regeneration of indigenous trees subsequent to the removal of plantation softwoods in Kibale National Park, Uganda. Counter to what would be expected if weeding released trees from competition, we found no difference in the total number of stems or in the stems greater than 10 cm diameter at breast height between control and weeded plots; there were more stems greater than 1 cm diameter at breast height in the control plots. For species found in both control and weeded plots, the maximum size of individuals did not differ. At the end of the study, 61 species were found in the control plots and 43 species were found in the weeded plots, and in both types of plots the three most abundant species were the same. The number of species and stems classified as early or middle successional species did not differ between weeded and control plots. The fact that weeding did not promote regeneration of indigenous trees after the removal of plantation trees illustrates the importance of evaluating and field‐testing potential management options.     

Restoration of morphological and functional integrity in the regenerating eye of the giant African land snail Achatina fulica

To determine whether vision returns to its original state following eye removal in Achatina fulica, light and electron microscope examinations, electrophysiological recordings and behavioural tests were carried out on the regenerating snails. Reparative morphogenesis can result in the restoration of the peripheral sense organ even in the absence of complete regrowth of the tentacle, but it can also lead to the formation of aberrant regenerates. We found that anatomically and ultrastructurally the eyes of the ‘most normal’ regenerates were basically the same as the original eyes. Under normal conditions each eye is composed of a principal and an accessory eye, both sharing a common cornea. The only difference between regenerated and native eyes is the smaller size of the former, as a result of a reduced number of retinal cells. Electroretinographic responses revealed that the molecular mechanism of phototransduction is restored, in principle, but that flicker fusion frequency in the regenerated eye is significantly lower than in the normal eye. The directional movement to a visual stimulus (a black stripe of 45° width) had not completely recovered even 6 months after amputation. This suggests that the central projections of the optic nerve had not become fully re‐established at the time of testing.     

Plant regeneration from protoplasts of the monocotyledonous Haworthia magnifica v. Poelln.

Calli were initiated from flower buds, gynoecia and inflorescence segments of Haworthia magnifica v. Poelln. and subcultured on solid medium. Two liquid culture steps were necessary to prepare the calli for the isolation of protoplasts capable of sustained cell divisions. Plants were regenerated from protoplast-derived calli. The influence of both the osmolality of the culture media and exudates on the viability of protoplasts and protoplast-derived cell colonies is briefly discussed.