Desert locusts (Schistocerca gregaria) feed with self-sharpening, scissor-like mandibles.

The mandibles of the desert locust Schistocerca gregaria (Forsskål, 1775) are digger-shovel-shaped mouthparts that are part of the locust's exoskeleton formed by the insect cuticle. The cuticle is a polymer-fibre composite, which supports, encases and protects the entire body. Mandibles experience heavy loading and wear due to direct contact with hard and abrasive food, just like teeth, their mineralized analogues in vertebrates. With dual-energy X-ray tomography, we image well-defined regions of zinc (Zn)-enriched cuticle at the mandible cutting edges and quantify the Zn concentrations in these regions. Zn is known to increase stiffness, hardness and wear resistance of the otherwise purely polymeric insect cuticle. In S. gregaria, the position of the Zn-enriched cutting-edge regions relative to one another suggests that the mandibles form a scissor-like cutting tool, which sharpens itself as the mouthparts shear past one another during feeding. Comparing the architecture of these purely polymeric mandibles with the mineralized incisors of rodents, we find fundamental design differences in cutting-tool structure and performance. Locusts' scissors and rodents' carving knives perform different functions, because they act on food that differs significantly in properties and shape: softer, sheet-like material in the case of locusts and harder bulk material in the case of rodents.

RAB7 GTPases as coordinators of plant endomembrane traffic.

The ras gene from rat brain (RAB) family of small GTPases is highly conserved among eukaryotes and regulates endomembrane trafficking pathways. RAB7, in particular, has been linked to various processes involved in regulating endocytic and autophagic pathways. Plants have several copies of RAB7 proteins that reflect the intricacy of their endomembrane transport systems. RAB7 activity regulates different pathways of endomembrane trafficking in plants: (1) endocytic traffic to the vacuole; (2) biosynthetic traffic to the vacuole; and (3) recycling from the late endosome to the secretory pathway. During certain developmental and stress related processes another pathway becomes activated (4) autophagic trafficking towards the vacuole that is also regulated by RAB7. RAB7s carry out these functions by interacting with various effector proteins. Current research reveals many unexplored RAB7 functions in connection with stress responses. Thus, this review describes a comprehensive summary of current knowledge of plant RAB7's functions, discusses unresolved challenges, and recommends prospective future research directions.

Terebra steering in chalcidoid wasps.

Various chalcidoid wasps can actively steer their terebra (= ovipositor shaft) in diverse directions, despite the lack of terebral intrinsic musculature. To investigate the mechanisms of these bending and rotational movements, we combined microscopical and microtomographical techniques, together with videography, to analyse the musculoskeletal ovipositor system of the ectoparasitoid pteromalid wasp Lariophagus distinguendus (Förster, 1841) and the employment of its terebra during oviposition. The ovipositor consists of three pairs of valvulae, two pairs of valvifers and the female T9 (9th abdominal tergum). The paired 1st and the 2nd valvulae are interlocked via the olistheter system, which allows the three parts to slide longitudinally relative to each other, and form the terebra. The various ovipositor movements are actuated by a set of nine paired muscles, three of which (i.e. 1st valvifer-genital membrane muscle, ventral 2nd valvifer-venom gland reservoir muscle, T9-genital membrane muscle) are described here for the first time in chalcidoids. The anterior and posterior 2nd valvifer-2nd valvula muscles are adapted in function. (1) In the active probing position, they enable the wasps to pull the base of each of the longitudinally split and asymmetrically overlapping halves of the 2nd valvula that are fused at the apex dorsally, thus enabling lateral bending of the terebra. Concurrently, the 1st valvulae can be pro- and retracted regardless of this bending. (2) These muscles can also rotate the 2nd valvula and therefore the whole terebra at the basal articulation, allowing bending in various directions. The position of the terebra is anchored at the puncture site in hard substrates (in which drilling is extremely energy- and time-consuming). A freely steerable terebra increases the chance of contacting a potential host within a concealed cavity. The evolution of the ability actively to steer the terebra can be considered a key innovation that has putatively contributed to the acquisition of new hosts to a parasitoid's host range. Such shifts in host exploitation, each followed by rapid radiations, have probably aided the evolutionary success of Chalcidoidea (with more than 500,000 species estimated).

Morphology of the abdominal segmental glands and spinning behaviour of Stenus larvae (Coleoptera, Staphylinidae).

We focus on the morphology of the "segmental glands" and their openings in third instar Stenus larvae. The location of the openings was similar in both studied species, with paired rosette-like structures present on the head, all thoracic segments and abdominal segments 1-9. No such openings could be found on the antennae, the maxillary palps, the urogomphi, and the legs as suggested in some older publications. We presume that the glands up to abdominal segment 7 are "adhesive" glands. They are compound glandular units consisting of a secretory syncytium with a common reservoir and a canal cell. The common reservoir is connected through a single efferent duct with the opening of the gland. Glands of abdominal segments 8 and 9 show differences in their length, number of reservoirs, the orientation of the efferent canal, the inner structures of the gland openings towards the exterior and the shape and content of the secretion vesicles indicating that they are silk glands for cocoon building. The spinning behaviour has been observed during the building of the hatching and pupation retreats. The larva first attaches to the substrate with its pygopod, secretes silk droplets from silk gland openings and pulls out a silk filament from the tip of its urogomphi. Whereas L1 and L2 instars produce an open single-layered net, L3 build a closed bi-layered cocoon.

Mouthpart Ecomorphology and Predatory Behaviour in Selected Rove Beetles of the "Staphylinine Group" (Coleoptera: Staphylinidae: Staphylininae, Paederinae).

The representatives of the megadiverse rove beetle subfamilies Paederinae and Staphylininae (Coleoptera: Staphylinidae) are considered generalist predators, although their exact prey-capture behaviour and performance and possible links to mouthpart morphology have rarely been described. Here, we examine these relationships for selected species by SEM analyses of mouthparts and front legs and highspeed videography of prey-capture behaviour. We describe the observed behaviours and structural properties and quantify relationships between prey type, mouthpart morphology, and predatory performance based on morphometric measurements of both the shape and lever properties of the mandible. We show that the Staphylininae considered have morphological and behavioural properties generally associated with generalist predation and that the Paederinae considered display characteristics that are highly specialized on elusive prey such as Collembola. We found correlations between mandible shape and leverage, and body size and prey type. We report distinct prey-capture behaviours: the beetles use front legs and/or mandibles to attack prey, drag prey, or cage it between their legs. These strategies differ among species and situations. Overall, this exploratory study provides insights into the morphology and types of prey capture that must have played a major role in the evolution of these beetles.

Osteogenic Differentiation of Human Adipose-Derived Stem Cells Seeded on a Biomimetic Spongiosa-like Scaffold: Bone Morphogenetic Protein-2 Delivery by Overexpressing Fascia.

Human adipose-derived stem cells (hADSCs) have the capacity for osteogenic differentiation and, in combination with suitable biomaterials and growth factors, the regeneration of bone defects. In order to differentiate hADSCs into the osteogenic lineage, bone morphogenetic proteins (BMPs) have been proven to be highly effective, especially when expressed locally by route of gene transfer, providing a constant stimulus over an extended period of time. However, the creation of genetically modified hADSCs is laborious and time-consuming, which hinders clinical translation of the approach. Instead, expedited single-surgery gene therapy strategies must be developed. Therefore, in an in vitro experiment, we evaluated a novel growth factor delivery system, comprising adenoviral BMP-2 transduced fascia tissue in terms of BMP-2 release kinetics and osteogenic effects, on hADSCs seeded on an innovative biomimetic spongiosa-like scaffold. As compared to direct BMP-2 transduction of hADSCs or addition of recombinant BMP-2, overexpressing fascia provided a more uniform, constant level of BMP-2 over 30 days. Despite considerably higher BMP-2 peak levels in the comparison groups, delivery by overexpressing fascia led to a strong osteogenic response of hADSCs. The use of BMP-2 transduced fascia in combination with hADSCs may evolve into an expedited single-surgery gene transfer approach to bone repair.

Root vacuolar sequestration and suberization are prominent responses of Pistacia spp. rootstocks during salinity stress.

Understanding the mechanisms of stress tolerance in diverse species is needed to enhance crop performance under conditions such as high salinity. Plant roots, in particular in grafted agricultural crops, can function as a boundary against external stresses in order to maintain plant fitness. However, limited information exists for salinity stress responses of woody species and their rootstocks. Pistachio (Pistacia spp.) is a tree nut crop with relatively high salinity tolerance as well as high genetic heterogeneity. In this study, we used a microscopy-based approach to investigate the cellular and structural responses to salinity stress in the roots of two pistachio rootstocks, Pistacia integerrima (PGI) and a hybrid, P. atlantica x P. integerrima (UCB1). We analyzed root sections via fluorescence microscopy across a developmental gradient, defined by xylem development, for sodium localization and for cellular barrier differentiation via suberin deposition. Our cumulative data suggest that the salinity response in pistachio rootstock species is associated with both vacuolar sodium ion (Na+) sequestration in the root cortex and increased suberin deposition at apoplastic barriers. Furthermore, both vacuolar sequestration and suberin deposition correlate with the root developmental gradient. We observed a higher rate of Na+ vacuolar sequestration and reduced salt-induced leaf damage in UCB1 when compared to P. integerrima. In addition, UCB1 displayed higher basal levels of suberization, in both the exodermis and endodermis, compared to P. integerrima. This difference was enhanced after salinity stress. These cellular characteristics are phenotypes that can be taken into account during screening for sodium-mediated salinity tolerance in woody plant species.

Osteoinduction within adipose tissue fragments by heterodimeric bone morphogenetic Proteins-2/6 and -2/7 versus homodimeric bone morphogenetic protein-2: Therapeutic implications for bone regeneration.

BACKGROUND: Fragments of subcutaneous adipose tissue that have been genetically modified to express bone morphogenetic protein-2 (BMP-2) regenerate large segmental osseous lesions in rodents. Gene-activated adipose tissue can be implanted into osseous defects without prior cell extraction and cell culture. The present study aimed to explore whether the heterodimers BMP-2/6 or BMP-2/7 exceed the osteoinductive effect of BMP-2 on adipose tissue. METHODS: In an in vitro tissue culture system, freshly harvested rat subcutaneous adipose tissue was cultivated in the presence of either BMP-2 or BMP-2/6 or BMP-2/7 at a high (200 ng/ml) and low (50 ng/ml) concentration. Gene expression analysis as well as histological and immunohistochemical methods were applied to test for osteoinduction. RESULTS: A concentration of 200 ng/ml of homodimeric BMP-2 induced osteogenic differentiation most potently, showing more calcification and a higher expression level of bone markers than both concentrations of BMP-2/6 or -2/7. A concentration of 50 ng/ml of BMP-2 was a significantly stronger osteogenic inducer than both concentrations of BMP-2/6 and the low concentration of BMP-2/7. The most potent heterodimeric driver of osteoinduction was BMP-2/7 at a high concentration, demonstrating effects similar to those of BMP-2 at a low concentration. CONCLUSIONS: Homodimeric BMP-2 evoked osteoinduction within adipose tissue more potently and at a lower concentration than heterodimeric BMP-2/6 or BMP-2/7. This result agrees well with the fact that it might be easier to translate adipose grafts activated by homodimeric BMP-2 clinically. Preclinical in vivo gene transfer studies are necessary to confirm the results of the present study.

Structure and function of the stylets of hematophagous Triatominae (Hemiptera: Reduviidae), with special reference to Dipetalogaster maxima.

Kissing bugs (Hemiptera: Reduviidae: Triatominae) are able to bend their rod-like maxillae while searching for blood vessels in the tissue of their vertebrate hosts. Little is known about the working mechanisms of these bending movements and the distal opening of the food channel. We compared the morphological structure of the stylets (mandibles and maxillae) of four triatomine species and analyzed the feeding process of Dipetalogaster maxima (Uhler, 1894). The maxillae of triatomine bugs are interlocked by a tongue-and-groove system, allowing longitudinal sliding. While penetrating the host tissue, the animals perform rapid alternate back and forth movements of the maxillae. The resistance of the surrounding tissue pushes the asymmetric apex of the maxillae away from its straight path, i.e., if one individual maxilla is protracted alone, its tip curves inwards, and the other maxilla follows. Once a blood vessel is tapped, the spine-like tip of the left maxilla splays outwards. Apically, each of the maxillae features an abutment, the left one exhibiting a notch that presumably facilitates splaying. The mechanical interaction of the two maxillary abutments enables the distal opening of the food channel but might also support the movements of the maxillary bundle attributable to different bending moment distributions.

Investigation of Salt Tolerance Mechanisms Across a Root Developmental Gradient in Almond Rootstocks.

The intensive use of groundwater in agriculture under the current climate conditions leads to acceleration of soil salinization. Given that almond is a salt-sensitive crop, selection of salt-tolerant rootstocks can help maintain productivity under salinity stress. Selection for tolerant rootstocks at an early growth stage can reduce the investment of time and resources. However, salinity-sensitive markers and salinity tolerance mechanisms of almond species to assist this selection process are largely unknown. We established a microscopy-based approach to investigate mechanisms of stress tolerance in and identified cellular, root anatomical, and molecular traits associated with rootstocks exhibiting salt tolerance. We characterized three almond rootstocks: Empyrean-1 (E1), Controller-5 (C5), and Krymsk-86 (K86). Based on cellular and molecular evidence, our results show that E1 has a higher capacity for salt exclusion by a combination of upregulating ion transporter expression and enhanced deposition of suberin and lignin in the root apoplastic barriers, exodermis, and endodermis, in response to salt stress. Expression analyses revealed differential regulation of cation transporters, stress signaling, and biopolymer synthesis genes in the different rootstocks. This foundational study reveals the mechanisms of salinity tolerance in almond rootstocks from cellular and structural perspectives across a root developmental gradient and provides insights for future screens targeting stress response.

A comparison of tarsal morphology and traction force in the two burying beetles Nicrophorus nepalensis and Nicrophorus vespilloides (Coleoptera, Silphidae).

Our aim was to compare friction and traction forces between two burying beetle species of the genus Nicrophorus exhibiting different attachment abilities during climbing. Specifically, the interaction of adhesive hairs and claws during attachment with respect to various surface properties was investigated by using a 2 × 3 experimental design. Traction force was measured for two different surface energies (hydrophilic vs hydrophobic) varying in roughness from smooth to micro-rough to rough. Nanotribometric tests on single legs were also performed. The external morphology of the attachment devices investigated by scanning electron microscopy suggested higher intra-specific (intersexual) than inter-specific differences. Whereas differences between the two species in traction force were high on smooth surfaces, no differences could be detected between males and females within each species. With claws intact, both species showed the highest forces on rough surfaces, although N. nepalensis with clipped claws performed best on a smooth surface. However, N. nepalensis beetles outperformed N. vespilloides, which showed no differences between smooth and rough surfaces with clipped claws. Both species demonstrated poor traction forces on micro-rough surfaces. Results concerning the impact of surface polarity were inconclusive, whereas roughness more strongly affected the attachment performance in both species. Nanotribometric analyses of the fore tarsi performed on micro-rough and rough surfaces revealed higher friction in the proximal (pull) direction compared with the distal (push) direction. In these experiments, we detected neither differences in friction performance between the two species, nor clear trends concerning the influence of surface polarity. We conclude that the investigated morphological traits are not critical for the observed interspecific difference in attachment ability on smooth surfaces. Furthermore, interspecific differences in performance are only clear on smooth surfaces and vanish on micro-rough and rough surfaces. Our results suggest that even subtle differences in the adhesion-mediating secretion in closely related species might result in qualitative performance shifts.

Gene activated adipose tissue fragments as advanced autologous biomaterials for bone regeneration: osteogenic differentiation within the tissue and implications for clinical translation.

Cost-effective, expedited approaches for bone regeneration are urgently needed in an ageing population. Bone Morphogenetic Proteins (BMPs) stimulate osteogenesis but their efficacy is impeded by their short half-life. Delivery by genetically modified cells can overcome this problem. However, cell isolation and propagation represent significant obstacles for the translation into the clinic. Instead, complete gene activated fragments of adipose tissue hold great potential for bone repair. Here, using an in-vitro culture system, we investigated whether adenoviral transduction with human BMP-2 can promote osteogenic differentiation within adipose tissue fragments. Osteoinduction in adipose tissue fragments was evaluated by quantitative reverse transcriptase polymerase chain reaction, immunohistology and histomorphometry. BMP-2 transduced adipose tissue synthesized BMP-2 protein over 30 days peaking by day six, which significantly promoted osteogenic differentiation as indicated by increased calcium depositions, up-regulation of bone marker genes, and bone-related protein expression. Our results demonstrate that cells within adipose tissue fragments can differentiate osteogenically after BMP-2 transduction of cells on the surface of the adipose tissue. BMP-2 gene activated adipose tissue represents an advanced osteo-regenerative biomaterial that can actively contribute to osteogenesis and potentially enable the development of a novel, cost-effective, one-step surgical approach to bone repair without the need for cell isolation.

Osteoinduction within BMP-2 transduced muscle tissue fragments with and without a fascia layer: implications for bone tissue engineering.

Bone can be engineered in vivo by implantation of gene-activated muscle tissue fragments. This expedited approach may be further improved by use of muscle tissue with attached fascia. The aim of this in vitro study was to provide an in depth comparison of the osteogenic differentiation capacity of muscle alone and muscle with fascia after BMP-2 transduction. Skeletal muscle tissue from rats was cut into pieces with and without a fascia layer on the surface. Adenoviral BMP-2 or GFP vectors were used for transduction. Osteogenic differentiation within the tissue fragments was evaluated and compared by qRT-PCR, alizarin red S staining, histomorphometry and immunohistology. Transduction efficiency and level of transgene expression were higher for muscle with fascia than muscle alone. Transduction with BMP-2 led to a significant upregulation of bone marker genes, proteins, and calcium deposition in both groups. Interestingly, histological evaluation revealed that osteoinduction did not occur within the fascia layer itself. The upregulation of bone marker genes in muscle with fascia was significantly lower after 2 weeks but similar after 4 weeks of in vitro culture in comparison to muscle alone. The fascia layer led to higher transduction efficiency and enhanced BMP-2 expression. Despite fascia's lower capacity for osteogenic differentiation, muscle implants may benefit from the fascia layer by the improved ability to deliver BMP-2. The presented data may contribute to the development of a novel, cost-effective, single-surgery bone engineering technology and encourage the evaluation of the osteoregenerative potential of muscle with fascia in an animal model.

Bone morphogenetic protein-2 is a stronger inducer of osteogenesis within muscle tissue than heterodimeric bone morphogenetic protein-2/6 and -2/7: Implications for expedited gene-enhanced bone repair.

BACKGROUND: Bone morphogenetic protein (BMP)-2 gene-activated muscle tissue fragments can regenerate large bone defects in preclinical animal models. The use of tissue fragments instead of isolated cells expedites gene-enhanced tissue engineering and may increase the possibility of clinical translation. The present in vitro study investigated whether the osteoinductive effect of BMP-2 on muscle tissue fragments can be enhanced using the heterodimers BMP-2/6 or BMP-2/7. METHODS: Skeletal muscle tissue fragments from rats were cultured in vitro for up to 20 days in normal medium, osteogenic medium or osteogenic medium supplemented with either a low (50 ng/ml) or high (200 ng/ml) concentration of recombinant human BMP-2, BMP-2/6 or BMP-2/7. Osteoinduction was evaluated by a quantitative reverse transcriptase-polymerase chain reaction, Alizarin red S staining, immunohistology and histomorphometry. RESULTS: Interestingly, BMP-2 was a significantly stronger inducer of osteogenic differentiation within muscle tissue than both heterodimers. Even the low concentration of BMP-2 elicited significantly higher levels of calcium deposition, bone-specific gene expression and protein production than the high concentration of both heterodimers. At the high concentration, BMP-2/7 had a significantly stronger osteogenic effect on muscle than BMP-2/6. CONCLUSIONS: The homodimer BMP-2 induced osteoblastogenesis in muscle faster, at a lower concentration and with a higher potency than the heterodimers BMP-2/6 or BMP-2/7. The findings of this in vitro study encourage bone repair by muscle implants in combination with BMP-2 single growth factor delivery, which might be beneficial with respect to clinical translation.

Morphology of hindwing veins in the shield bug Graphosoma italicum (Heteroptera: Pentatomidae).

Light, fluorescence, and electron microscopy were applied to cross sections and -breakage and whole-mount preparations of the anterior hindwing vein of the shield bug Graphosoma italicum. These analyses were complemented by investigations of the basal part of the forewing Corium and Clavus. The integration of structural, histological, and fluorescence data revealed a complex arrangement of both rigid and elastic structures in the wall of wing veins and provided insights into the constitution of transition zones between rigid and elastic regions. Beneath the exocuticular layers, which are continuous with the dorsal and ventral cuticle of the wing membrane, the lumen of the veins is encompassed by a mesocuticular layer, an internal circular exocuticular layer, and an internal longitudinal endocuticular layer. Separate parallel lumina within the anterior longitudinal vein of the hindwing, arranged side-by-side rostro-caudally, suggest that several veins have fused in the phylogenetic context of vein reduction in the pentatomid hindwing. Gradual structural transition zones and resilin enrichment between sclerotized layers of the vein wall and along the edges of the claval furrow are interpreted as mechanical adaptations to enhance the reliability and durability of the mechanically stressed wing veins.

Migration of Mesenchymal Stem Cells of Bursal Tissue after Rotator Cuff Repair in Rats.

Purpose The purpose of this study is to verify migration of mesenchymal stem cells of bursal tissue into the healing site after rotator cuff repair in rats. Methods Fischer rats and green fluorescent protein (GFP)-transgenic rats were used. Bursal tissue from GFP rats was isolated and transplanted into tendon repair sites in Fischer rats. We examined the histology of the rotator cuff and the proportion of GFP-positive cells in the repaired rotator cuff 1, 3, and 6 weeks after surgery. Results Cell migration was observed during the third and sixth week after surgery. We also found mesenchymal stem cells and formed bursal cluster patterns in the repaired rotator cuff tendons. Conclusion Mesenchymal stem cells migrated from bursal tissue and infiltrated the repaired rotator cuff tendons. Clinical Relevance Mesenchymal stem cells from bursal tissue can contribute to the healing progress of the repaired rotator cuff.

Jumping mechanisms and performance in beetles. II. Weevils (Coleoptera: Curculionidae: Rhamphini).

We describe the kinematics and performance of the natural jump in the weevil Orchestes fagi (Fabricius, 1801) (Coleoptera: Curculionidae) and its jumping apparatus with underlying anatomy and functional morphology. In weevils, jumping is performed by the hind legs and involves the extension of the hind tibia. The principal structural elements of the jumping apparatus are (1) the femoro-tibial joint, (2) the metafemoral extensor tendon, (3) the extensor ligament, (4) the flexor ligament, (5) the tibial flexor sclerite and (6) the extensor and flexor muscles. The kinematic parameters of the jump (from minimum to maximum) are 530-1965 m s-2 (acceleration), 0.7-2.0 m s-1 (velocity), 1.5-3.0 ms (time to take-off), 0.3-4.4 µJ (kinetic energy) and 54-200 (g-force). The specific joint power as calculated for the femoro-tibial joint during the jumping movement is 0.97 W g-1. The full extension of the hind tibia during the jump was reached within up to 1.8-2.5 ms. The kinematic parameters, the specific joint power and the time for the full extension of the hind tibia suggest that the jump is performed via a catapult mechanism with an input of elastic strain energy. A resilin-bearing elastic extensor ligament that connects the extensor tendon and the tibial base is considered to be the structure that accumulates the elastic strain energy for the jump. According to our functional model, the extensor ligament is loaded by the contraction of the extensor muscle, while the co-contraction of the antagonistic extensor and flexor muscles prevents the early extension of the tibia. This is attributable to the leverage factors of the femoro-tibial joint providing a mechanical advantage for the flexor muscles over the extensor muscles in the fully flexed position. The release of the accumulated energy is performed by the rapid relaxation of the flexor muscles resulting in the fast extension of the hind tibia propelling the body into air.

Macro-SICM: A Scanning Ion Conductance Microscope for Large-Range Imaging.

The scanning ion conductance microscope (SICM) is a versatile, high-resolution imaging technique that uses an electrolyte-filled nanopipet as a probe. Its noncontact imaging principle makes the SICM uniquely suited for the investigation of soft and delicate surface structures in a liquid environment. The SICM has found an ever-increasing number of applications in chemistry, physics, and biology. However, a drawback of conventional SICMs is their relatively small scan range (typically 100 µm × 100 µm in the lateral and 10 µm in the vertical direction). We have developed a Macro-SICM with an exceedingly large scan range of 25 mm × 25 mm in the lateral and 0.25 mm in the vertical direction. We demonstrate the high versatility of the Macro-SICM by imaging at different length scales: from centimeters (fingerprint, coin) to millimeters (bovine tongue tissue, insect wing) to micrometers (cellular extensions). We applied the Macro-SICM to the study of collective cell migration in epithelial wound healing.

Recent advances in gene-enhanced bone tissue engineering.

The loss of bone tissue represents a critical clinical condition that is frequently faced by surgeons. Substantial progress has been made in the area of bone research, providing insight into the biology of bone under physiological and pathological conditions, as well as tools for the stimulation of bone regeneration. The present review discusses recent advances in the field of gene-enhanced bone tissue engineering. Gene transfer strategies have emerged as highly effective tissue engineering approaches for supporting the repair of the musculoskeletal system. By contrast to treatment with recombinant proteins, genetically engineered cells can release growth factors at the site of injury over extended periods of time. Of particular interest are the expedited technologies that can be applied during a single surgical procedure in a cost-effective manner, allowing translation from bench to bedside. Several promising methods based on the intra-operative genetic manipulation of autologous cells or tissue fragments have been developed in preclinical studies. Moreover, gene therapy for bone regeneration has entered the clinical stage with clinical trials for the repair of alveolar bone. Current trends in gene-enhanced bone engineering are also discussed with respect to the movement of the field towards expedited, translational approaches. It is possible that gene-enhanced bone tissue engineering will become a clinical reality within the next few years.

Gene-activated tissue grafts for sustained bone morphogenetic protein-2 delivery and bone engineering: Is muscle with fascia superior to muscle and fat?

Previously, we have presented an expedited strategy for sustained delivery of bone morphogenetic protein-2 (BMP-2) to bone lesions based on the implantation of gene-activated fat and muscle fragments. The aim of the present in vitro experiments was to evaluate the potential of muscle with fascia as a BMP-2 delivering osteo-regenerative implant in comparison to fat tissue and muscle alone. Subcutaneous fat, muscle, and muscle with fascia were harvested from Fischer 344 rats. The tissues were cut into small pieces and cultured for up to 90 days after direct transduction with adenoviral BMP-2 or green fluorescence protein vectors. Different vector doses were applied, and proliferation, long-term BMP-2 production, and osteogenic differentiation of the 3 different tissues were investigated in vitro. Muscle with fascia produced the largest amounts of BMP-2. Expression of the transgene was detected for up to 90 days. Proliferation was reduced with increased vector doses. Muscle with fascia showed a higher potential for osteogenic differentiation than fat, but it was not improved as compared to muscle alone. A dose of 4 × 108 plaque forming units of the adenoviral BMP-2 vector appeared to be the optimal dose for transduction of muscle with fascia. Because muscle with fascia produced higher amounts of BMP-2 as compared to muscle alone or fat tissue grafts, showing a high potential for osteogenic differentiation, it might represent an improved osteo-regenerative implant facilitating endogenous repair. Future studies should investigate the effect of muscle with fascia transduced with 4 × 108 plaque forming units on bone healing in vivo.