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Biomedical subjects

Mauro Alini

Publications and source records attributed to Mauro Alini.

At least 19 recordsLinked to original sources
PubMed2007

The long-term effects of ovariectomy on bone metabolism in sheep.

Osteoporosis and associated fractures are major public health concerns, and as such require appropriate large animal models to further our understanding of this disease. Although sheep appear to be an ideal model with which to study bone loss caused by estrogen depletion, limited data are available concerning the long-term effect of ovariectomy on bone in sheep. The goal of the present study was to observe the ovariectomy-induced changes in bone mass, structure, and metabolism in sheep over a period of 18 months. Six ewes were ovariectomized (OVX) and compared to an age-matched control group by analyzing bone mineral density, trabecular structure, biochemical markers of bone formation and resorption, and plasma estrogen levels. Bone loss (13%, P < 0.01) occurred during the first 4 months after surgery, then stabilized and returned to pre-OVX levels for the remainder of the study. Trabecular architecture was also altered and tended toward osteopenia with recovery to baseline values. Markers of bone formation and resorption were elevated up to 6 months postovariectomy, after which time levels returned to baseline values. Although estradiol measurements demonstrated a clear decline following surgical ovariectomy, levels returned to normal after 6 months. Therefore, the detrimental effect of ovariectomy on sheep bone metabolism seems to be reversible, with normal bone parameters being reestablished within 6 months after surgery. These data seem to indicate that the sheep is not an appropriate model for human postmenopausal osteoporosis.

Age Factors↗
PubMed2006

Seasonal changes in bone metabolism in sheep.

There is a great need for animal models of osteoporosis and sheep are a suitable large animal that meets most requirements. Since it is known that bone mass in humans responds to seasonal changes, this study investigated natural bone metabolism in sheep in order to better define the sheep as a model for osteoporosis. Bone mineral density (BMD), trabecular structure, biochemical markers of bone formation and resorption and estrogen were analysed over a period of 18 months. The lowest BMDs, measured by peripheral quantitative computed tomography (pQCT), were observed during winter. Thereafter, a 5.1% increase in BMD was observed during spring and summer (P<0.05). Bone resorption markers showed a variable pattern, with higher values in spring compared to autumn (P<0.001). The physiological estrus phase during autumn was detected by serum estrogen levels. The findings show that it is necessary to take seasonal variations into account if sheep are used to establish an animal model for osteoporosis.

Amino Acids↗
PubMed2006

An in vitro organ culturing system for intervertebral disc explants with vertebral endplates: a feasibility study with ovine caudal discs.

STUDY DESIGN: Whole ovine caudal intervertebral discs with vertebral endplates were cultured under uniaxial diurnal loading for 7 days. OBJECTIVES: To establish and characterize an organ culture system for intervertebral discs, in which disc cells may be "maintained" in their native three-dimensional environment under load. SUMMARY OF BACKGROUND DATA: In vitro culturing of entire discs with preserved biologic and structural integrity would be a useful model to study the effects of nutrition and mechanical loading. METHODS: To maintain endplate permeability, sheep were systemically anticoagulated before death and their caudal vasculature was evacuated with saline postmortem. The first 4 caudal discs were explanted with their adjacent endplates and cultured in bioreactors under uniaxial diurnal loading (0.2 MPa for 8 hours and 0.8 MPa for 16 hours) for 4 or 7 days. Solute transport into the center of the disc was measured after 4 days of culture using a low molecular weight fluorescent marker. Cell viability, glycosaminoglycan synthesis rate, and gene expression profile were measured after 7 days of culture and compared with fresh tissue. RESULTS: Fluorescent images showed that solutes could diffuse into the disc under both static and diurnal loading, but penetration through the endplate increased with diurnal loading. Cell viability and glycosaminoglycan synthesis rates remained unchanged after 7 days of culture. Expression of catabolic genes was significantly up-regulated, whereas anabolic genes tended to be down-regulated after 7 days. CONCLUSIONS: With this novel preparation and culturing technique, endplate permeability could be maintained, which allowed culturing of intact disc explants with endplates for up to 7 days.

Animals↗
PubMed2006

Human endothelial cells inhibit BMSC differentiation into mature osteoblasts in vitro by interfering with osterix expression.

It is well accepted, that there is communication between osteoblasts and endothelial cells. However, the influence of endothelial cells on the differentiation of bone precursors into mature osteoblasts is not yet well understood. We therefore studied the effect of human umbilical vein endothelial cells (HUVEC) on human bone marrow stromal cell (BMSC) differentiation towards an osteoblastic phenotype by culturing them in two different types of HUVEC-BMSC cultures (indirect contact, HUVEC-conditioned medium). Typical bone markers (45Ca incorporation, ALP activity, and gene expression of collagen 1 (COL1), osteonectin (ON), matrix metalloproteinase 13 (MMP-13), bone morphogenetic protein 2 (BMP-2)) and transcription factors (Runx2, osterix (OSX)) were evaluated at different time points. The effect of stimulating HUVEC with vascular endothelial growth factor (VEGF) before co-cultures with BMSC was also evaluated. As expected, BMSC in osteogenic medium (OM) (with dexamethasone) differentiated towards the osteoblastic phenotype, as measured by increased matrix mineralization, high ALP activity, and elevated expression of specific osteoblastic marker genes, when compared to BMSC in non-OM. HUVEC reversibly inhibited osteoblastic differentiation of BMSC in OM, independent of the co-culture type. When HUVEC were removed from the cultures, BMSC differentiated into osteoblasts, albeit at a faster rate than BMSC that were never co-cultured with HUVEC. Stimulation of HUVEC with VEGF before co-culture enhanced the inhibitory effect of HUVEC on BMSC differentiation. This inhibitory effect was connected to a reversible suppression of OSX gene expression and was specific to endothelial cells. We conclude that HUVEC can inhibit dexamethasone-induced BMSC differentiation into osteoblasts in vitro, by interfering with OSX expression, thereby arresting BMSC differentiation at a preosteoblastic stage.

Anti-Inflammatory Agents↗
PubMed2006

Quantitative magnetic resonance imaging of enzymatically induced degradation of the nucleus pulposus of intervertebral discs.

STUDY DESIGN: The structural integrity of the nucleus pulposus (NP) of intervertebral discs was targeted by enzyme-specific degradations to correlate their effects to the magnetic resonance (MR) signal. OBJECTIVE: To develop quantitative MR imaging as an accurate and noninvasive diagnostic tool to better understand and treat disc degeneration. SUMMARY OF BACKGROUND DATA: Quantitative MR analysis has been previously shown to reflect not only the disc matrix composition, but also the structural integrity of the disc matrix. Further work is required to identify the contribution of the structural integrity versus the matrix composition to the MR signal. METHODS: The bovine coccygeal NPs were injected with either enzyme or buffer, incubated at 37 degrees C as static, unloaded and closed 3-disc segments, and analyzed by a 1.5-Tesla MR scanner to measure MR parameters. RESULTS: Collagenase degradation of the NP significantly decreased the relaxation times, slightly decreased the magnetization transfer ratio, and slightly increased the apparent diffusion coefficient. Targeting the proteoglycan and/or hyaluronan integrity by trypsin and hyaluronidase did not significantly affect the MR parameters, except for an increase in the apparent diffusion coefficient of the disc after trypsin treatment. CONCLUSIONS: Our results demonstrate that changes in the structural integrity of matrix proteins can be assessed by quantitative MR.

Animals↗
PubMed2006

In vitro organ culture of the bovine intervertebral disc: effects of vertebral endplate and potential for mechanobiology studies.

STUDY DESIGN: Whole bovine coccygeal discs were cultured under static load, with or without vertebral endplates (VEPs), and assessed for cell viability, biochemical stability, biosynthetic activity, and biosynthetic responsiveness to changes in mechanical load. OBJECTIVES: To assess the effects of VEPs on biochemical and cellular stability of disc cells during in vitro culture of large disc explants. To determine whether cultured discs could respond to mechanical perturbation. SUMMARY OF BACKGROUND DATA: Previous methods for culturing the intervertebral disc have focused on rabbit and rat discs, but the small size of these discs limits the relevance of these culture systems to the human condition. Bovine coccygeal discs have similar dimensions to the human lumbar disc (i.e., similar size and nominal stresses), but long-term culture of these discs has not been reported. METHODS: Bovine coccygeal discs were harvested with or without VEPs, cultured under static load (5 kg, approximately 0.25 MPa, in situ swelling pressure) for up to 1 week, and evaluated for changes in hydration, glycosaminoglycan content, cell viability, and biosynthetic activity. Additionally, the biochemical and biosynthetic response of discs cultured without VEP to increasing the load to a 20-kg (approximately 1 MPa, the estimated stress in human lumbar disc during heavy lifting) static load for 6 hours was assessed. RESULTS: During the first 24 hours, culturing discs with endplates was moderately better with regards to maintaining in situ anulus hydration and nucleus glycosaminoglycan levels. The endplates, however, obstructed media flow to the disc, resulting in a marked decrease in cell viability after 1 week of culture. Nucleus pulposus cell viability was maintained in discs cultured without endplates, but there was a significant drop in biosynthetic activity within 2 days of culture. Despite this drop, the disc cells in the discs without VEP remained biosynthetically responsive to changes in mechanical loading. CONCLUSIONS: It is possible to maintain cell viability and the biosynthetic responsiveness of large discs for up to 1 week in vitro when the discs are cultured under static load and without VEP.

Animals↗
PubMed2006

Biodegradable elastomeric polyurethane membranes as chondrocyte carriers for cartilage repair.

Autologous chondrocyte implantation in combination with an autologous periosteal patch has become a clinically accepted procedure for the treatment of articular cartilage defects. The use of periosteum has, however, several drawbacks. We have been able to fabricate thin elastomeric biodegradable polyurethane (PU) membranes that may possibly have an application as a tissue-engineered substitute for the periosteal patch. Three types of membranes varying in pore size and surface texture were used as substrates for bovine chondrocytes in culture. The membranes, marked as P-I, P-II, and P-R, had average pore sizes of 10 to 20 microm, 40 to 60 microm, and less than 5 microm, respectively. A poly(L/DL-lactide) 80/ 20% micro-porous membrane (PLA) with an average pore size in the range of 10 to 70 microm was used as a control. There was no difference in the cell proliferation profile among the 4 membranes. In terms of proteoglycan and collagen production, P-I, P-R, and PLA performed similarly to one another. The rate of matrix production appears to be greater in the PU membranes than in the PLA membrane in the first 10 days, although by day 30, the PLA membrane had caught up. In all comparisons, the performance of P-II lagged behind those of the other materials. In conclusion, this preliminary study supports the potential use of this novel group of PUs as a periosteal flap substitute or perhaps as a chondrocyte carrier for matrix-assisted chondrocyte implantation and related techniques. Further studies will be necessary to better define their role in clinical applications for cartilage repair.

Absorbable Implants↗
PubMed2006

Leptin inhibits bone formation not only in rodents, but also in sheep.

UNLABELLED: This study examines the effect of long-term ICV administration of leptin in ewes. We found that central application significantly decreased osteoblast activity as measured by serum analysis as well as by histomorphometry, resulting in decreased trabecular bone volume. These data provide additional evidence that bone formation and therefore bone remodeling is at least in part centrally controlled. INTRODUCTION: Genetic studies in mice have identified leptin as a potent inhibitor of bone formation acting through the central nervous system and unraveled the central nature of bone mass control and its disorders. Although these studies have radically enhanced our understanding of skeletal physiology because they have established a hypothalamic regulation of bone remodeling through the sympathetic nervous system, controversy remains about the physiological relevance of these observations because leptin's effect on bone after intracerebroventricular (ICV) application has only been shown in mice. To address whether leptin has a role in regulating bone mass beyond rodents, we treated ewes with long-term ICV application of leptin and analyzed the bone phenotype after a treatment period of 3 months. MATERIALS AND METHODS: Three groups of corriedale sheep were compared: (1) control entire (control), (2) ovariectomy (OVX) and ICV application of cerebrospinal fluid (CSF); and (iii) OVX and ICV application of leptin (leptin). Analysis included histomorphometric characterization of iliac crest, spine and femur by histology and biomechanical testing and measurement of bone turnover parameters in serum and urine. RESULTS: Central application of leptin decreased bone formation by 70% and mineralizing surface (MS/BS, 39.4 +/- 3.3% versus 16.1 +/- 2.1%) significantly (p < 0.01). Whereas OVX increased osteoclast indices and urinary cross-lap excretion by two and three times, respectively, serum parameters of osteoblast activity were significantly reduced by ICV application of leptin (p < 0.01). Consequently, ewes treated with leptin were osteopenic (iliac crest BV/TV entire, 22.7 +/- 1.3%; CSF, 18.9 +/- 2.4%; leptin, 12.4 +/- 2.6%), whereas bone torsional failure load reflecting the cortex of the tibia was not yet changed after 3 months of treatment (p < 0.01). CONCLUSIONS: Taken together, these data suggest that leptin controls bone formation after ICV application, leading to reduction of trabecular bone mass in sheep. Most importantly, however, they show that the central regulation of bone formation is not limited to rodents, but is also found in large animals, providing further evidence that bone remodeling in vertebrates is centrally controlled.

Animals↗
PubMed2006

Effects of mechanical loading on intervertebral disc metabolism in vivo.

The overall goal of this work is to define more clearly which mechanical loading conditions are associated with accelerated disc degeneration. This article briefly reviews recent studies describing the effects of mechanical loading on the metabolism of intervertebral disc cells and defines hypothetical models that provide a framework for quantitative relationships between mechanical loading and disc-cell metabolism. Disc cells respond to mechanical loading in a manner that depends on loading magnitude, frequency, and duration. On the basis of the current data, four models have been proposed to describe the effects of continuous loading on cellular metabolism: (1) on/off response, in which messenger ribonucleic acid (mRNA) transcription remains altered for the duration of loading; (2) maintenance, characterized by an initial change in mRNA levels with return to baseline levels; (3) adaptation, in which mRNA transcription is altered and remains at a new steady state; and (4) no response. In addition, five hypothetical mechanisms that describe the long-term consequences of these metabolic changes on disc-remodeling are presented. The transient nature of gene expression along with enzyme activation/inhibition is associated with changes at the protein level. The hypothetical models presented provide a framework for obtaining quantitative relationships between mechanical loading, gene expression, and changes at the compositional level; however, additional factors, such as regulatory mechanisms, must also be considered when describing disc-remodeling. A more quantitative relationship between mechanical loading effects and the metabolic response of the disc will contribute to injury prevention, facilitate more effective rehabilitation treatments, and help realize the potential of biologic and tissue engineering approaches toward disc repair.

Animals↗
PubMed2005

The potential of chitosan-based gels containing intervertebral disc cells for nucleus pulposus supplementation.

The suitability of chitosan-based hydrogels as scaffolds for the encapsulation of intervertebral disc (IVD) cells and the accumulation of a functional extracellular matrix mimicking that of the nucleus pulposus (NP) was investigated. The specific hypothesis under study was that the cationic chitosan would form an ideal environment in which large quantities of newly synthesized anionic proteoglycan could be entrapped. Indeed, all the formulations of cell-seeded chitosan hydrogels, studied under in vitro culture conditions, showed that the majority of proteoglycan produced by encapsulated NP cells was retained within the gel rather than released into the culture medium. This was not always the case when annulus fibrosus cells were encapsulated, as unlike the nucleus cells the annulus cells often did not survive when cultured in chitosan. The results support the concept that chitosan may be a suitable scaffold for cell-based supplementation to help restore the function of the NP during the early stages of IVD degeneration.

Animals↗
PubMed2005

The effects of short-term load duration on anabolic and catabolic gene expression in the rat tail intervertebral disc.

The goal of this study was to determine the time-dependent response of the intervertebral disc cells to in vivo dynamic compression. Forty-seven skeletally mature Wistar rats (>12 months old) were instrumented with an Ilizarov-type device spanning caudal disc 8-9. Using a load magnitude (1 MPa) and frequency (1.0 Hz) that were previously shown to significantly alter mRNA levels in the disc, the effects of 0.5 and 4 h of loading were investigated and compared to a sham group and our previous 2 h results. Annulus and nucleus tissue of loaded (c8-9) and internal control discs (c6-7 and c10-11) were separately analyzed by real-time RT-PCR for levels of mRNA coding for various anabolic (collagen-1A1, collagen-2A1, aggrecan) and catabolic (MMP-3, MMP-13, ADAMTs-4) proteins. In the annulus, mRNA levels increased for Collagen types I & II, and MMP 3 & 13 with increasing load duration. In contrast, the nucleus had the largest increases in aggrecan, ADAMTs-4, MMP-3 and MMP-13 after 2 h of loading, with aggrecan and MMP-13 mRNA levels returning to control values after 4 h of loading. Taken in context with our previous studies, we conclude that intervertebral disc cells from the nucleus and annulus have distinct responses to dynamic mechanical compression in vivo with sensitivity to compression magnitude, frequency and duration.

ADAM Proteins↗
PubMed2005

2004 Young Investigator Award Winner: vertebral endplate marrow contact channel occlusions and intervertebral disc degeneration.

STUDY DESIGN: Intervertebral disc degeneration was evaluated by morphologic appearance, magnetic resonance imaging, and by biochemical matrix composition. Caliber and distribution of openings of the adjacent vertebral osseous endplates were measured. OBJECTIVES: Correlation between occlusion of vertebral endplate openings and intervertebral disc degeneration was quantified. SUMMARY OF BACKGROUND DATA: Calcifications of vertebral endplates with disease and age have suggested insufficient nutrition as a mechanism for intervertebral disc degeneration. It has been proposed that occlusion of endplate openings, which contain vascular sources for the disc, may limit the transport of nutrients, leading to disc degeneration. METHODS: Fresh magnetic resonance images from 39 human lumbar discs were scored. Sectioned discs with endplates were morphologically graded. Samples of nuclear and anular regions were evaluated for proteoglycan and collagen contents. Backlight microscopic images of 4 endplate regions were obtained, and caliber and distribution of endplate openings for each disc were measured. Analysis of variance regression models were used to assess correlation between endplate openings and disc degeneration. RESULTS: The decrease in opening density significantly correlated to morphologic degeneration grade, best for openings with 20 to 50 im equivalent diameter and in the nuclear region. Although the density of 20 to 50 im openings also significantly indirectly correlated to age, it was not as strong as the correlation to degeneration grade. Opening density was also significantly correlated to proteoglycan content in all regions. However, all other biochemical parameters as well as the T2 intensity score showed only weak or no correlation to opening density. CONCLUSIONS: A high indirect correlation between the density of openings in the osseous endplate (particularly of the size of the capillary buds) and the morphologic degeneration grade of the disc support the hypothesis that occlusion of these openings may deprive the cells of nutrients, leading to insufficient maintenance of the extracellular matrix and disc degeneration.

Adult↗
PubMed2004

Correlation of radiographic and MRI parameters to morphological and biochemical assessment of intervertebral disc degeneration.

UNLABELLED: Degenerative disc disease (DDD) is a common finding in MRI scans and X-rays. However, their correlation to morphological and biochemical changes is not well established. In this study, radiological and MRI parameters of DDD were assessed and compared with morphological and biochemical findings of disc degeneration. Thirty-nine human lumbar discs (L1-S1), age 19-86 years, were harvested from eight cadavers. Within 48 h postmortem, MRIs in various spin-echo sequences and biplanar radiographs of intact spines were obtained. Individual discs with endplates were then sectioned in the mid-sagittal plane and graded according to the morphological appearance. Samples from the nucleus of each disc were harvested for biochemical analysis including water and proteoglycan contents. On MRIs, T2-signal intensity, Modic changes, disc extension beyond the interspace (DEBIT), nucleus pulposus shape, annular tears, osteophytes and endplate integrity were graded. On radiographs, an independent observer classified the parameters disc height, endplate sclerosis, osteophytes, Schmorl's nodes, intradiscal calcifications and endplate shape. General linear-regression models were used for statistical analysis. Backward elimination with a 10% significance cut-off level was used to identify the most significant parameters, which then were summed to create composite scores for radiography, MRI and the combination of both methods. The grading was performed by three observers, and a reliability analysis using Cronbach's alpha model was used to control interobserver agreement. The three radiographic parameters height-loss, osteophytes and intradiscal calcifications correlated significantly with the morphological degree of degeneration (p<0.001, R2=642). Significant differences of even one morphological grade could also be differentiated in the composite radiological score (p<0.05), except at the extremes between grades 1 and 2 and grades 4 and 5. All MRI parameters correlated significantly with the morphological grade (p<0.05); however Modic changes, T2-intensity and osteophytes accounted for 83% of the variation in the data. T2-signal intensity correlated significantly with H2O and proteoglycan content (p<0.001), and was best for detecting highly degenerated discs. Regression showed that the combined score was better correlated with the morphological grade (p<0.001, R2=775) than either the composite radiographic (p<0.001, R2=642) or composite MRI (p<0.001, R2=696) alone. Based on the combined score, a backwards elimination of the regression was performed, in which the parameters Modic changes, and T2-intensity loss (MRI) as well as calcifications (X-ray) accounted for 87% of the variability. The interobserver validation showed a high correlation for all three scores (Cronbach's alpha values ranging from 0.95 to 0.97). CONCLUSION: Selective imaging parameters and a newly created scoring scheme were found to correlate with disc degeneration as determined in a morphological manner. Surprisingly, radiographic parameters were able to distinguish different stages of degeneration, whereas MRI could only detect advanced stages of disc degeneration. We conclude that X-rays may remain a cost-effective, non-invasive in vivo-grading method to detect early disc degeneration, and, combined with MRI, correlate best with morphological and biochemical assessment of disc degeneration.

Adult↗
PubMed2004

Apparent diffusion coefficient of intervertebral discs related to matrix composition and integrity.

While tremendous work has been performed to characterize degenerative disc disease through gross morphologic, biochemical, and histologic grading schemes, the development of an accurate and noninvasive diagnostic tool is required to objectively detect changes in the matrix with aging and disc degeneration. In the present study, quantitative magnetic resonance was used to determine if the quality of the nutritional supply to the intervertebral disc at various ages and levels of degeneration could be assessed through measurement of the apparent diffusion coefficients (ADCs). Modifications of the nucleus pulposus matrix content, specifically of water and glycosaminoglycan contents, with age and disc degeneration, were reflected in correlating changes in the ADCs. From unforced stepwise linear regression analyses, relations were established showing that decreases in glycosaminoglycan or water contents in the nucleus pulposus resulted in direct decreases in the ADCs. Relations obtained for the ADCs of the nucleus pulposus were direction dependent, in conformity with the anisotropic diffusion in the intervertebral discs. Changes in matrix integrity, as evidenced by the percentage of denatured collagen, were also detected in the nucleus pulposus with a low positive correlation to the ADC along the height of the disc and an inverse statistically significant regression to the ADC along the anterior to posterior axis of the disc. Correlations between the matrix content and integrity of the annulus fibrosus and its ADCs were not as evident, with only the ADC in the lateral direction of the disc of the anterior annulus fibrosus able to reflect changes in matrix content. The information obtained by the ADCs, particularly of the nucleus pulposus, can potentially be used in combination with quantitative T1, T2, and MT parameters to noninvasively obtain a quantitative assessment of the disc matrix composition and structural integrity.

Adolescent↗
PubMed2004

Anabolic and catabolic mRNA levels of the intervertebral disc vary with the magnitude and frequency of in vivo dynamic compression.

The goal of this study was to characterize the anabolic and catabolic mRNA response of the disc to dynamic loading to determine if variations in the magnitude and/or frequency of loading could elicit different cellular responses. Sixty-eight Wistar rats were instrumented with an Ilizarov-type device spanning caudal disc 8-9. Seventy-two hours after surgery, animals were anesthetized and loaded at either 1 or 0.2 MPa at a frequency of 1, 0.2 or 0.01 Hz for 2 h (6 groups). The surgical control (Sham) animals underwent anesthesia with no loading. Loaded (c8-9) and internal-control discs (c6-7 and c10-11) were dissected and annulus and nucleus tissue were separately analyzed by real-time RT-PCR for levels of anabolic (collagen-1A1, collagen-2A1, aggrecan) and catabolic (MMP-3, MMP-13, ADAMTs-4) mRNA. In the nucleus, a frequency-dependent response was seen at 1 MPa with anabolic genes stimulated at 0.01 Hz and catabolic genes at 1 Hz. In the annulus all frequencies resulted in significant up-regulation of catabolic mRNA at 1 MPa loading. In general loading at 0.2 MPa or 0.2 Hz had little effect on gene expression. The results suggest that gene expression of the annulus appears to be more dependent on the magnitude of applied stress, while the nucleus is both magnitude- and frequency-dependent.

ADAM Proteins↗
PubMed2003

Chondrocytes seeded onto poly (L/DL-lactide) 80%/20% porous scaffolds: a biochemical evaluation.

Repair of articular cartilage remains a clinical and scientific challenge. The use of implantable chondrocyte-seeded constructs offers a promising treatment option. The present study is focused on the culture of chondrocytes onto 3-D poly (L-/DL-lactide) 80%/20% porous scaffolds. Scaffolds were seeded with bovine articular chondrocytes and cultured for up to 42 days. After an initial decrease, although not significant, the DNA content of the constructs remained constant over the culture time. A progressive increase in glycosaminoglycans and collagen was observed in the constructs during the culture period. However, a significant release of matrix molecules into the culture medium also was noticeable. On the transcriptional level, aggrecan mRNA expression was highest on day 14 and gradually decreased thereafter. Procollagen I mRNA expression constantly increased whereas procollagen II mRNA expression decreased after day 14, indicating that cells were dedifferentiating. Histologic analysis of the constructs demonstrated the deposition of a proteoglycan-rich extracellular matrix by day 42 of culture. The present data demonstrate that poly (L-/DL-lactide) 80%/20% porous scaffolds support the development of a cartilage-like extracellular matrix. The limitations of the system are the diffusion of large amounts of matrix molecules into the culture medium and the dedifferentiation of the chondrocytes with prolonged time in culture.

Animals↗
PubMed2003

Effects of immobilization and dynamic compression on intervertebral disc cell gene expression in vivo.

STUDY DESIGN: An in vivo analysis of the intervertebral disc's cellular response to dynamic compression and immobilization was performed using a rat-tail model. OBJECTIVE: To assess the effects of immobilization and short-term dynamic compression on intervertebral disc cell expression of anabolic and catabolic genes. SUMMARY OF BACKGROUND DATA: Static compressive loads applied in vivo alter the composition of the disc matrix and cell viability in a dose-dependent manner. The effects of in vivo dynamic compression, which is a more physiologic load, and reported risk factor for low back pain have not been investigated. METHODS: An Ilizarov-type device was implanted on the rat tail and used to determine the effects from 72 hours of immobilization (n = 6), 2 hours of dynamic compression (1 MPa/0.2 Hz) (n = 8), and the coupled effect of immobilization followed by compression (n = 8). Real-time reverse transcription-polymerase chain reaction was used to measure changes in anabolic and catabolic gene levels relative to both internal control subjects and a sham-operated group (n = 7). RESULTS: Immobilization and dynamic compression affect anabolic and catabolic genes, with an overall downregulation of types 1 and 2 collagen and upregulation of aggrecanase, collagenase, and stromelysin in the anulus. The effects of immobilization and compression appear to be additive for collagen types 1 and 2 in the anulus, but not in the nucleus, and not for catabolic genes. CONCLUSIONS: Short-duration dynamic compression and immobilization alter gene expression in the rat disc. In studying the response of the disc to loading, it is necessary to look at both anabolic and catabolic pathways, and to consider strain history.

Aggrecans↗