Biomedical subjects

Robert M Pilliar

Publications and source records attributed to Robert M Pilliar.

At least 19 recordsLinked to original sources

PubMed2006

Differential regulation of matrix degrading enzymes in a TNFalpha-induced model of nucleus pulposus tissue degeneration.

Intervertebral disc degeneration occurs commonly and is linked to persistent back pain and the development of disc herniation. The mechanisms responsible for tissue catabolism have not yet been fully elucidated. Previously we characterized an in vitro model of TNFalpha-induced nucleus pulposus degeneration, which demonstrates decreased expression of matrix macromolecules, increased expression of matrix degrading enzymes, and the activation of aggrecanase-mediated proteoglycan degradation [Seguin, C.A., Pilliar, R.M., Roughley, P.J., and Kandel, R.A. 2005. Tumor necrosis factor-alpha modulates matrix production and catabolism in nucleus pulposus tissue. Spine 30: 1940-1948]. This study explores the intracellular pathways activated during TNFalpha-induced matrix degradation. We demonstrate that in nucleus pulposus cells, the p38 and JNK pathways regulate induction of MMP-1 and -3; p38, JNK, and NF-kappaB regulate the induction of MMP-13; and ERK regulates the up-regulation of MT1-MMP mRNA in response to TNFalpha. Induction of ADAMTS-4 and -5 mRNA occurred downstream of NF-kappaB activation. Depletion of tissue proteoglycans was mediated by ERK and NF-kappaB-dependent "aggrecanase" activity, suggesting MT1-MMP and ADAMTS-4 and -5 as effectors of TNFalpha-induced tissue catabolism.

ADAM Proteins↗

PubMed2006

Heterotopic bone formation around sintered porous-surfaced Ti-6Al-4V implants coated with native bone morphogenetic proteins.

PURPOSE: Coating endosseous dental implants with growth factors such as bone morphogenetic proteins (BMPs) may be one way to accelerate and/or enhance the quality of osseointegration. The purpose of this study was to investigate in the murine muscle pouch model whether sintered porous-surfaced titanium alloy implants coated with BMPs would lead to heterotopic bone formation around and within the implant surface geometry. MATERIALS: Porous-surfaced dental implants were coated with partially purified native human BMPs, with or without a carrier of Poloxamer 407 (BASF Corp., Parsippany, NJ), placed in gelatin capsules and implanted into the hindquarter muscles of mice. Mice were euthanized after 28 days. Sections of retrieved specimens were subsequently prepared for morphometric analysis of bone formation using backscatter electron microscopic images. RESULTS: Human BMPs, either with or without the carrier of Poloxamer 407, led to bone formation within and outside of the sintered porous implant surface. When the sintered implant surface region was subdivided into inner and outer halves, similar levels of bone ingrowth and contact were seen in the 2 halves. Evidence of bone formation to the depth of the solid implant core (i.e., the deepest level possible) also was seen. DISCUSSION AND CONCLUSIONS: Sintered porous-surfaced dental implants can be used as substrate for partially purified BMPs in the murine muscle pouch model. With the addition of these osteoinductive factors, the porous implant surface supported bone formation within the surface porosity provided, in some instances, all the way to the solid implant core. The addition of growth factors to a sintered porous surface may be an efficient method for altering locally the healing sequence and quality of bone associated with osseointegration of bone-interfacing implants.

Alloys↗

PubMed2005

Formation of a nucleus pulposus-cartilage endplate construct in vitro.

Intervertebral disc (IVD) degeneration is a common problem and treatment options for persistent symptomatic disease are limited. Tissue engineering is being explored for its ability to reconstitute the functional components of the IVD. The purpose of this study was to determine whether it was possible to form in vitro a triphasic construct consisting of nucleus pulposus (NP), cartilage endplate (CEP), and a porous calcium polyphosphate (CPP) bone substitute. Bovine articular chondrocytes were placed on the top surface of a porous CPP construct and allowed to form cartilage in vitro. Nucleus pulposus cells were then placed onto the in vitro-formed hyaline cartilage. At 24 h scanning electron microscopy demonstrated that the NP cells maintained their rounded morphology, similar to NP cells placed directly on porous CPP. At 8 weeks histological examination of the triphasic constructs by light microscopy showed that a continuous layer of NP tissue had formed and was fused to the underlying cartilage tissue, which itself was integrated with the porous CPP. The incorporation of the cartilage layer was beneficial to the construct by improving tissue attachment to the CPP, as demonstrated by increased peak load and increased energy required for failure during shear loading when compared to a biphasic construct composed of nucleus pulposus-bone substitute only. This study demonstrates that it is possible to generate a multi-component construct with the incorporation of a CEP-like layer resulting in improved bone substitute-to-IVD tissue interface characteristics.

Animals↗

PubMed2005

Tumor necrosis factor-alpha modulates matrix production and catabolism in nucleus pulposus tissue.

STUDY DESIGN: This study examines changes in the production of extracellular matrix molecules as well as the induction of tissue degradation in in vitro formed nucleus pulposus (NP) tissues following incubation with tumor necrosis factor (TNF)alpha. OBJECTIVE: To characterize the response of NP cells to TNF-alpha, a proinflammatory cytokine present in herniated NP tissues. SUMMARY OF BACKGROUND DATA: TNF-alpha is a proinflammatory cytokine expressed by NP cells of degenerate intervertebral discs. It is implicated in the pain associated with disc herniation, although its role in intervertebral disc degeneration remains poorly understood. METHODS: In vitro formed NP tissues were treated with TNF-alpha (up to 50 ng/mL) over 48 hours. Tissues were assessed for histologic appearance, proteoglycan and collagen contents, as well as proteoglycan and collagen synthesis. Reverse transcriptase polymerase chain reaction was used to determine the effect of TNF-alpha on NP cell gene expression. Proteoglycan degradation was assessed by immunoblot analysis. RESULTS: At doses of 1-5 ng/mL, TNF-alpha induced multiple cellular responses, including: decreased expression of both aggrecan and type II collagen genes; decreases in the accumulation and overall synthesis of aggrecan and collagen; increased expression of MMP-1, MMP-3, MMP-13, ADAM-TS4, and ADAM-TS5; and induction of ADAM-TS dependent proteoglycan degradation. Within 48 hours, these cellular responses resulted in NP tissue with only 25% of its original proteoglycan content. CONCLUSIONS: Because low levels of TNF-alpha, comparable to those present physiologically, induced NP tissue degradation, this suggests that TNF-alpha may contribute to the degenerative changes that occur in disc disease.

ADAM Proteins↗

PubMed2005

Influence of anionic monomer content on the biodegradation and toxicity of polyvinyl-urethane carbonate-ceramic interpenetrating phase composites.

The objective of this study was to characterize a series of anionic biodegradable polymer resins for their compatibility in a biological environment, comparing them with respect to the influence of ionic function on enzyme catalyzed biodegradation when the polymers were incorporated into a porous calcium polyphosphate (CPP) 3-D structure to form an interpenetrating phase composite (IPC). The swelling behavior of the polymers was investigated by immersing the cured polymer resins in growth media at 37 degrees C. In vitro cytotoxicity of the polymer resins was assessed using a HeLa cell line. Cell viability increased when the amount of low molecular weight monomer was minimized. Despite observing that the addition of carboxylic acid groups into the polymer resin chains contributed to an improvement of the chemical bonding between the polymer and the CPP, the addition of high ionic content into the resin led to the greatest loss of bending strength for the samples incubated in phosphate buffer and cholesterol esterase enzyme solutions, when compared to their as made state. The increased degradation for the higher ionic component materials and their loss of physical strength was attributed to enhanced hydrolysis within the materials and by water transport deep within the composites, via the anionic components of the resin. The findings indicated that the introduction of anionic content must be optimized to promote increased mechanical performance for the CPP, balancing the features of polymer CPP bonding versus polymer swelling and cytotoxicity.

Anions↗

PubMed2005

Peri-implant bone response to orthodontic loading: Part 1. A histomorphometric study of the effects of implant surface design.

INTRODUCTION: Bone response to orthodontic loading was compared histomorphometrically around 2 different types of osseointegrated implants (porous surfaced and machined threaded) to determine their suitability for orthodontic anchorage. METHODS: Five beagles each received 3 implants of each design in contralateral mandibular locations. After a 6-week initial healing period, abutments were placed, and, 1 week later, the 2 mesial implants on each side were orthodontically loaded for 22 weeks. All implants remained osseointegrated throughout orthodontic loading except for 1 threaded implant that loosened. Light miscroscopy and back-scattered scanning electron microscopy were used to compare responses around the 2 implant designs. RESULTS: Porous-surfaced implants had higher marginal bone levels (P +/- .025) and less relative implant displacement than threaded implants. CONCLUSIONS: Differences in implant surface design can lead to differences in peri-implant bone height and bone-to-implant contact. Porous-surfaced implants might be successful as orthodontic anchorage units.

Alveolar Process↗

PubMed2005

Peri-implant bone response to orthodontic loading: Part 2. Implant surface geometry and its effect on regional bone remodeling.

INTRODUCTION: Bone response to orthodontic loading was compared around 2 different types of osseointegrated implants (porous surfaced and machined threaded) to determine the effect of implant surface geometry on regional bone remodeling. METHODS: Five beagles each received 3 implants of each design in contralateral mandibular extraction sites. After a 6-week initial healing period, abutments were placed, and, 1 week later, the 2 mesial implants on each side were orthodontically loaded for 22 weeks. All implants remained osseointegrated throughout orthodontic loading except for 1 threaded implant that loosened. Back-scattered scanning electron microscopy and fluorochrome bone labeling techniques were used to compare responses around the 2 types of implants. RESULTS: The loaded, porous-surfaced implants had significantly higher marginal bone levels and greater bone-to-implant contact than did the machined-threaded implants. CONCLUSIONS: Significant differences in peri-implant bone remodeling and bone formation in response to controlled orthodontic loading were observed for the 2 implant designs. Short, porous-surfaced implants might be more effective for orthodontic applications than machine-threaded implants.

Alveolar Process↗

PubMed2005

Evaluating interface strength of calcium phosphate sol-gel-derived thin films to Ti6Al4V substrate.

The interface shear strength of Ca-P thin films applied to Ti6Al4V substrates have been evaluated in this study using a substrate straining method--a shear lag model. The Ca-P films were synthesized using sol-gel methods from either an inorganic or organic precursor solution. Strong interface bonding was demonstrated for both film types. The films were identified as non-stoichiometric hydroxyapatite but with different Ca/P ratios. The Ca-P films were 1-1.5 microm thick and testing and analysis using the shear lag approach revealed a shear strength of approximately 347 and 280 MPa for Inorganic and Organic Route-formed films, respectively. Overall, the exceptional mechanical properties of Ca-P/Ti6Al4V system along with the inherent advantages of sol-gel processing support continued studies to utilize this technology for bone-interfacing implant surface modification.

Adhesiveness↗

PubMed2005

Cementless implant fixation--toward improved reliability.

Cementless implants offer the advantage of fixation by direct bone-to-implant osseointegration, thereby avoiding the use of a synthetic intermediary material (such as acrylic bone cement) of limited mechanical strength. Successful osseointegration, however, depends on several conditions being satisfied during the peri-implant bone healing period, including the need for limited early loading resulting in minimal relative movement at the implant-bone interface. Sintered porous- and plasma spray-coated implants represent the most common cementless orthopedic implants in current clinical use, although novel cast structures also are being investigated. All stand to benefit from surface modifications currently being explored to enhance osteoconductive or osteoinductive characteristics of the implants. The faster osseointegration that such modified surface designs potentially might offer would result in more reliable and convenient (from the patient perspective) cementless implants. Encouraging results of early animal-based studies exploring such modifications have been reported.

Alloys↗

PubMed2005

Osteotome sinus elevation and simultaneous placement of porous-surfaced dental implants: a morphometric study in rabbits.

The objective was to establish a model in rabbits in which to study the healing events associated with localized indirect osteotome-mediated maxillary sinus floor elevation in conjunction with simultaneous placement of sintered porous-surfaced dental implants. On one side of the maxilla of each of 28 rabbits, a sintered porous-surfaced titanium alloy press-fit implant was placed without the use of a bone graft material, while on the collateral side an implant was placed after first adding Bio-Oss graft particles to the osteotomy. Specimens were retrieved for morphometric assessment of bone contact and bone ingrowth of the porous implant surface after 2, 4, 6 and 8 weeks of healing. All implants became osseointegrated by bone ingrowth into the porous implant surface. While the addition of graft particles did not result in a statistically significant increase in the parameters measured, a trend for greater bone contact and particularly bone ingrowth at the apices of the implants was seen as healing time increased. The rabbit maxillary sinus can be used to study healing following placement of sintered porous-surfaced dental implants using the indirect sinus elevation procedure.

Alloys↗

PubMed2004

Tissue engineered nucleus pulposus tissue formed on a porous calcium polyphosphate substrate.

STUDY DESIGN: This study describes the formation of nucleus pulposus tissue using a novel tissue engineering approach. OBJECTIVES: To determine if a construct composed of nucleus pulposus tissue on the surface of a calcium polyphosphate substrate could be formed in vitro with properties similar to native nucleus pulposus tissue. SUMMARY OF BACKGROUND DATA: There is no optimal treatment for the persistent pain associated with intervertebral disc degeneration. Disc replacement using artificial intervertebral discs has met with some success, and biologic transplantation is limited by the availability of donor tissues. METHODS: Nucleus pulposus cells were isolated from bovine caudal intervertebral discs. Cells were seeded at high density on the upper surface of a porous bone substitute material (calcium polyphosphate) and maintained up to 6 weeks in culture. In vitro formed tissue was compared to native nucleus pulposus for histologic appearance, biochemical composition (tissue cellularity, proteoglycan and collagen accumulation), and compressive mechanical properties. RESULTS: When maintained on the surface of a three-dimensional substrate, nucleus pulposus cells formed a continuous layer of tissue with a proteoglycan content equivalent to the native tissue. Although collagen accumulation attained only 26% than that of the native tissue, there was no difference in tissue stiffness, viscosity, or weight-bearing capacity of the in vitro formed tissue when compared with the native tissue. CONCLUSION: Nucleus pulposus-like tissue formed in vitro on the surface of a calcium polyphosphate substrate resembles the native tissue in terms of proteoglycan content and compressive mechanical properties. These studies are the first step toward developing a functional spinal unit in vitro.

Animals↗

PubMed2003

Synthesis and characterization of a novel polymer-ceramic system for biodegradable composite applications.

The objective of this study was to develop a biodegradable polymer resin that could be used for the fabrication of an interpenetrating phase composite (IPC) made of porous calcium polyphosphate (CPP) and an organic polymer resin. The resin was synthesized from a polycarbonate-based divinyl oligomer and monomers containing ionic groups. The physical and chemical properties of the polymer resin and polycarbonate-based divinyl oligomer were characterized by gel permeation chromatography, Fourier transform infrared spectroscopy, and swelling studies. The in vitro degradation of the polymer resins was assessed using cholesterol esterase in a buffer solution at 37 degrees C for 3 weeks. Scanning electron microscopy of the degraded samples indicated that the hydrolysis of the resin was catalyzed by the enzyme. The relative interfacial shear strength between the polymer resin and the CPP ceramic was studied using a microbond test. The addition of ionic groups into the polymer resin chains appeared to improve the chemical bonding between the polymer and the CPP. Preliminary mechanical properties of the IPC were investigated by determining bending strength using a three point bending test. The data showed a sevenfold increase in strength over that of the monolithic CPP, and the addition of more ionic groups into the resin led to a higher bending strength for the newly formed CPP/polycarbonate resin system. Sample cross sections of the IPC examined using scanning electron microscopy suggested that the resin had infiltrated almost all of the pores of the CPP. The results of this study indicate that the IPC could potentially be used for fabricating novel biodegradable load-bearing implants.

Biocompatible Materials↗

PubMed2003

The use of specific chondrocyte populations to modulate the properties of tissue-engineered cartilage.

Tissue engineering of articular cartilage is a promising alternative to the conventional approaches for cartilage repair. However, recent attempts to develop articular cartilage in vitro have proven to be difficult. The tissue formed in vitro may not accumulate enough extracellular matrix, and the resulting mechanical properties are only a fraction of the native tissue. We investigated whether using specific populations of chondrocytes would improve the properties of the cartilaginous tissue that was generated in vitro. Full-thickness (FT), mid-and-deep zone (MD), and deep-zone (DEEP) chondrocytes were isolated, placed on the surface of porous ceramic substrates and maintained in culture for eight weeks. Tissue developed from DEEP chondrocytes was thicker (FT: 0.94+/-0.03, MD: 0.88+/-0.04, DEEP: 2.4+/-0.1 mm) and had accumulated larger amounts of extracellular matrix (FT: 1.61+/-0.05, MD: 1.5+/-0.1, DEEP: 3.8+/-0.2 mg dry weight) than the tissues formed by the FT and MD chondrocytes. The tissue formed by the FT chondrocytes accumulated the greatest amount of collagen (FT: 211+/-14, MD: 185+/-8, DEEP: 178+/-5 microg/mg dry weight) whereas the tissue formed by the MD chondrocytes accumulated significantly more proteoglycans (FT: 198+/-10, MD: 265+/-10, DEEP: 215+/-5 microg/mg dry weight). Interestingly, MD chondrocytes produced tissue that had compressive mechanical properties up to four times greater than the cartilaginous tissues formed by cells from either the FT or DEEP of cartilage. Thus, a combined population of intermediate and DEEP chondrocytes might be more suitable for the tissue engineering of articular cartilage.

Animals↗

PubMed2003

Long-term intermittent shear deformation improves the quality of cartilaginous tissue formed in vitro.

The formation of cartilaginous tissue in vitro is a promising alternative to repair damaged articular cartilage. However, recent attempts to tissue-engineer articular cartilage that has similar properties to the native tissue have proven to be difficult. The in vitro-formed cartilaginous tissue typically has a similar proteoglycan content to native cartilage, but has a reduced collagen content and only a fraction of the mechanical properties. In this study, we investigated whether the intermittent application of cyclic shearing forces during tissue formation would improve the tissue quality. Chondrocyte cultures were stimulated at a 2% shear strain amplitude at a frequency of 1 Hz for 400 cycles every 2nd day. At one week, both collagen and proteoglycan synthesis increased (23+/-6% and 20+/-6%, respectively) over the unstimulated, static controls. At four weeks, an increased amount of tissue formed (stimulated: 1.85+/-0.08, unstimulated: 1.58+/-0.07 mg dry wt.). This tissue contained approximately 40% more collagen (stimulated: 511+/-23, unstimulated: 367+/-24 microg/construct) and 35% more proteoglycans (stimulated: 376+/-21, unstimulated: 279+/-26 microg/construct). Tissues that formed in the presence of shearing forces also displayed a 3-fold increase in compressive load-bearing capacity (stimulated: 16+/-5, unstimulated: 5+/-1 kPa max. equilibrium stress) and a 6-fold increase in stiffness (stimulated: 112+/-36, unstimulated: 20+/-6 kPa max. equilibrium modulus) compared to the static controls. These results demonstrate that intermittent application of dynamic shearing forces over a four-week period improves the quality of cartilaginous tissue formed in vitro. Interestingly, low amplitudes of shear stimulation for short periods of time (6 min of stimulation applied every 2nd day) produced these changes.

Animals↗

PubMed2003

Effect of biomechanical conditioning on cartilaginous tissue formation in vitro.

BACKGROUND: Although tissue engineering of articular cartilage is a promising approach for cartilage repair, it has been difficult to develop cartilaginous tissue in vitro that mimics the properties of native cartilage. Isolated chondrocytes grown in culture typically do not accumulate enough extracellular matrix, and the generated tissue possesses only a fraction of the mechanical properties of native cartilage. One potential explanation for this might be that the cells are grown in an environment that lacks the mechanical stimuli to which the chondrocytes are exposed in vivo. In this study, we compared the long-term effects of both dynamic compressive and shearing forces on cartilaginous tissue formation in vitro. METHODS: Bovine articular chondrocytes were grown on the surface of porous ceramic substrates and were maintained under static, free-swelling conditions for a period of four weeks. Cultures were then subjected to six minutes of mechanical stimulation every other day, in either compression or shear, for an additional four-week period. RESULTS: Cartilaginous tissues cultured in the presence of intermittent compression or shear were significantly thicker (p < 0.05) and had accumulated more extracellular matrix (p < 0.01) compared with the unstimulated controls. However, when normalized by the wet weight of the tissue, cultures stimulated in the presence of shearing forces contained more proteoglycans and collagen compared with compression-stimulated cultures. These cultures also displayed the largest increase in mechanical properties, with a threefold increase in equilibrium stress and a fivefold increase in equilibrium modulus. CONCLUSIONS AND CLINICAL RELEVANCE: The results of this study demonstrate that a brief application of mechanical forces applied periodically over a long duration can improve the quality of cartilaginous tissue formed in vitro. However, the changes in tissue composition and mechanical properties were dependent on the specific mode of the applied mechanical forces, with shear stimulation eliciting the greater effect. This finding suggests that chondrocytes may respond differently to different modes of applied forces.

Animals↗

PubMed2002

Characterization of cartilagenous tissue formed on calcium polyphosphate substrates in vitro.

Successful joint resurfacing by tissue-engineered cartilage has been limited, in part, by an inability to secure the implant to bone. To overcome this, we have developed the methodology to form a cartilage implant in vitro consisting of a layer of cartilagenous tissue overlying a porous, biodegradable calcium polyphosphate (CPP) substrate. As bone will grow into the CPP after implantation, it will result in anchorage of the cartilage. In this study, the cartilagenous tissue formed in vitro after 8 weeks in culture was characterized and compared to native articular cartilage. Light microscopic examination of histological sections showed that there was a continuous layer of cartilagenous tissue on, and integrated with the subsurface of, the CPP substrate. The in vitro-formed tissue achieved a similar thickness to native articular cartilage (mean +/- SEM: in vitro = 0.94 +/- 0.03 mm; ex vivo = 1.03 +/- 0.01 mm). The cells in the in vitro-formed tissue synthesized large proteoglycans (Kav +/- SEM: in vitro = 0.27 +/- 0.01; ex vivo = 0.27 +/- 0.01) and type II collagen similar to the chondrocytes in the ex-vivo cartilage. The in vitro-formed tissue had a similar amount of proteoglycan (GAG microg/mg dry wt.: in vitro = 198 +/- 10; ex vivo = 201 +/- 13) but less collagen than the native cartilage (hydroxyproline microg/mg dry wt.: in vitro = 21 +/- 1; ex vivo = 70 +/- 8). The in vitro-formed tissue had only about 3% of the load-bearing capacity and stiffness of the native articular cartilage, determined from unconfined mechanical compression testing. Although low, this was within the range of properties reported by others for tissue-engineered cartilage. It is possible that the limited load-bearing capacity is the result of the low collagen content and further studies are required to identify the conditions that will increase collagen synthesis.

Animals↗

PubMed2002

Attachment, spreading, and matrix formation by human gingival fibroblasts on porous-structured titanium alloy and calcium polyphosphate substrates.

Porous calcium polyphosphate (CPP) structures represent promising resorbable implant systems that can promote anchorage to connective tissues. Previous studies focused on chondrocyte interactions with CPP, but there are limited data on interactions of soft connective tissue cells with these materials. We studied attachment, spreading, and matrix formation by human gingival fibroblasts when cultured on amorphous and crystalline CPP. Comparison with porous Ti6Al4V substrates of similar volume percent, porosity, and pore size distribution provided evaluations of fibroblast interactions with rapid, moderate, and nonbiodegradable systems, respectively. Cells were incubated on substrates in medium containing ascorbic acid and evaluated at 3, 24, 48, 72, and 96 h after plating. Attached cell counts, cytoplasmic actin filament area, and immunostained extracellular type 1 collagen or fibronectin were quantified by morphometric analyses using epifluorescence microscopy. Cell morphology and substrate interactions were evaluated by scanning electron microscopy. Spreading, attachment, and matrix production were similar for both CPP substrates. In contrast, titanium alloy substrates exhibited threefold more attachment and twofold more spreading than CPP substrates. The area per cell of immunostained extracellular collagen and fibronectin was similar for the three different substrates. The results indicate that the crystallinity and, hence, degradation rate of CPP substrates does not substantially affect the interactions of fibroblasts with CPP materials but that compared with titanium alloy substrates, spreading and attachment are inhibited.

Alloys↗

PubMedDate not supplied

Long-term intermittent compressive stimulation improves the composition and mechanical properties of tissue-engineered cartilage.

Tissue engineering of articular cartilage is a promising alternative for cartilage repair. However, it has been difficult to develop tissue in vitro that mimicks native cartilage. Cartilaginous tissue formed in vitro does not accumulate enough extracellular matrix, is deficient in collagen, and possesses only a fraction of the mechanical properties of native cartilage. In this study, we investigated whether long-term intermittent compressive stimulation would improve the quality of the generated tissue. Chondrocyte cultures were established on the surface of porous calcium polyphosphate substrates and allowed to form cartilaginous tissue. In vitro-formed tissues were subjected to different stimulation protocols for 1 week. The optimal mechanical stimulation parameters identified in this short-term study were then applied to the cultures for up to 4 weeks. Mechanical stimulation applied at a 5% compressive amplitude at a frequency of 1 Hz for 400 cycles every second day resulted in the greatest increase in collagen synthesis (37 +/- 9% over control) while not significantly affecting proteoglycan synthesis (2 +/- 8% over control). This condition, applied to the chondrocyte cultures for 4 weeks, resulted in a significant increase in the amount of tissue that formed (stimulated, 2.4 +/- 0.2 mg dry wt; unstimulated, 1.61 +/- 0.08 mg dry wt). Stimulated tissues contained approximately 40% more collagen (stimulated, 590 +/- 58 microg; unstimulated, 420 +/- 42 microg), and 30% more proteoglycans (stimulated, 393 +/- 34 microg; unstimulated, 302 +/- 32 microg) as well as displaying a 2- to 3-fold increase in compressive mechanical properties (maximal equilibrium stress: stimulated, 10 +/- 1 kPa; unstimulated, 5 +/- 1 kPa; maximal equilibrium modulus: stimulated, 80 +/- 23 kPa; unstimulated, 24 +/- 6 kPa). The results of this study demonstrate that intermittent mechanical stimulation can increase collagen synthesis and, when applied over a 4-week period, can accelerate extracellular matrix accumulation as well as improve the material properties of the developed tissue. Interestingly, only short periods of mechanical stimulation (6 min every second day) were needed to affect the quality of cartilaginous tissue formed in vitro.

Adaptation, Physiological↗