Biomedical subjects

J Glowacki

Publications and source records attributed to J Glowacki.

At least 37 records · Page 2Linked to original sources

PubMed2001

Engineered cartilage, bone, joints, and menisci. Potential for temporomandibular joint reconstruction.

Reconstruction of many musculoskeletal structures can be accomplished by bone grafting and implantation of prostheses. Alternate approaches are needed, however, for repair of complex structures such as articular cartilage surfaces and the temporomandibular meniscus and joint. Tissue engineering, either cell-free or cell-based, offers promise because of recent advances in materials research and in our knowledge of the cellular and molecular mechanisms of tissue repair. There are three considerations in designing a construct for engineered tissue: the source of cells, if any; the nature of the carrier or scaffold; and use, if any, of genes, factors, or adjuvants. Autogenous cells, often expanded in vitro, have been useful for cartilage tissue engineering. Precursor/progenitor cells are advantageous for bone tissue. There are many natural and synthetic resorbable materials with good biocompatibility and tissue compatibility that can be modified to have the porosity and mechanical properties needed for specific applications. The scaffolds can also be modified to provide biological signals to augment repair and integration.

Animals↗

PubMed2000

Age-related decline in osteoprotegerin expression by human bone marrow cells cultured in three-dimensional collagen sponges.

With advancing age, an increase in bone resorption relative to bone formation results in bone loss. Bone marrow stromal cells and their products support osteoclastogenesis from hematopoietic progenitors. Another of their products, osteoprotegerin (OPG), blocks the osteoclast-stimulatory effects of OPG ligand. We tested the hypothesis that with advancing age there is a decrease in OPG expression by human bone marrow cells. Bone marrow cells were obtained from 18 subjects (age range 38-84 years). Expression of mRNA transcripts of OPG was assessed by quantitative competitive RT-PCR. Median number of OPG transcripts in the younger group was 0. 3 zetptomoles (range 0.01 to 1.30) and was higher than in the older group's median of 0.06 (range 0 to 0.5; p < 0.05). The decline in the expression of OPG with age may increase the capacity of stromal/osteoblast cells to support osteoclastogenesis.

Adult↗

PubMed2000

Effects of age on serum dehydroepiandrosterone sulfate, IGF-I, and IL-6 levels in women.

Data from animal and in vitro studies suggest that the growth-promoting effects of the adrenal androgen dehydroepiandrosterone sulfate (DHEAS) may be mediated by stimulation of insulin-like growth factor-I (IGF-I) and/or inhibition of interleukin 6 (IL-6), a cytokine mediator of bone resorption. This study tests the hypotheses that there are effects of age on serum DHEAS, IGF-I, and IL-6 levels, and that levels of IGF-I and IL-6 are related to DHEAS levels. The study included 102 women: 27 premenopausal and 75 postmenopausal, including 35 postmenopausal women with osteoporosis, as defined by bone mineral density scores by dual X-ray energy absorptiometry. DHEAS levels decreased significantly with age (r = -0.52, P < 0.0001) and IGF-I levels decreased significantly with age (r = -0.49, P < 0.0001). IL-6 levels increased significantly with age (r = 0.36, P = 0.008). IGF-I was positively correlated to DHEAS levels (r = 0.43, P < 0. 0001, n = 102) and IL-6 levels were negatively correlated to DHEAS levels (r = -0.32, P = 0.021, n = 54). Levels of DHEAS and IGF-I were correlated with T scores of the spine and some hip sites. In a multiple variable model to predict DHEAS, age was an important predictor (P < 0.001), but osteoporosis status, IGF-I, and IL-6 were not. The median DHEAS level was lower in the postmenopausal osteoporotic women (67 microg/dl, n = 35) than in the nonosteoporotic postmenopausal women (106.3 microg/dl, n = 40, P = 0. 03), but this was not significant after correction for age. Age accounted for 32% of the variance in DHEAS levels. In summary, DHEAS levels decreased with age and had a positive association with IGF-I levels and a negative association with IL-6 levels. DHEA deficiency may contribute to age-related bone loss through anabolic (IGF-I) and anti-osteolytic (IL-6) mechanisms.

Absorptiometry, Photon↗

PubMed2000

Effects of latency and rate on bone formation in a porcine mandibular distraction model.

PURPOSE: Long treatment protocols currently limit the application of distraction osteogenesis (DO). The purpose of this study was to develop a porcine model for DO and to define the effects of latency and distraction rate on bone formation. MATERIALS AND METHODS: Distractors were placed through submandibular incisions. For analysis of latency, mandibular osteotomies were distracted after 0 (n = 3) or 4 (n = 2) days at a rate of 1 mm/d (7 days) with 14 days fixation. For analysis of rate, osteotomies were distracted at 1 mm (n = 4), 2 mm (n = 4), or 4 mm (n = 4) per day to produce a 12-mm gap with 24 days fixation. DO wounds were assessed in vivo with bimanual palpation to detect mobility across the gap. Harvested specimens were evaluated by gross appearance and bimanual palpation. Standardized radiographic techniques were used to estimate bone density. RESULTS: During DO, animals showed normal activity. There were no infections, and no distractors were removed prematurely. Clinical and radiographic evaluation of the groups that underwent distraction after 0- or 4-day latency showed equivalent healing. In the comparison of distraction rates, stability was greatest in the group distracted at 1 mm/d. CONCLUSIONS: A porcine model for mandibular DO has been developed in which mandibular lengthening was successfully performed without latency and at a rate of 1 mm/d. The relationships among latency, gap size, rate, and duration of fixation are poorly understood and can be defined with relevant animal models.

Animals↗

PubMed1999

Occult vitamin D deficiency in postmenopausal US women with acute hip fracture.

CONTEXT: Low vitamin D levels may contribute to hip fractures in women, although limited data are available on vitamin D levels in US women admitted with acute hip fractures. OBJECTIVE: To determine whether postmenopausal women with hip fractures have low vitamin D and high parathyroid hormone levels compared with nonosteoporotic and osteoporotic women admitted for elective joint replacement. DESIGN: Comparative case series conducted between January 1995 and June 1998. SETTING AND PATIENTS: Ninety-eight postmenopausal community-dwelling women with no secondary causes of bone loss admitted for hip replacement, of whom 30 women had acute hip fractures and 68 women were admitted for elective joint replacement. Of the women admitted for elective joint replacement, 17 had osteoporosis and 51 did not. Women with comorbid conditions or who were taking medications that affect bone density and turnover were excluded. MAIN OUTCOME MEASURES: Primary measures were levels of vitamin D and parathyroid hormone; secondary measures were body composition and markers of bone turnover. RESULTS: Women with hip fractures had lower levels of 25-hydroxyvitamin D than women without osteoporosis admitted for elective joint replacement (P = .02) and than women with osteoporosis admitted for elective joint replacement (P = .01) (medians, 32.4, 49.9, and 55.0 nmol/L, respectively; comparisons adjusted for age and estrogen intake). Parathyroid hormone levels were higher in women with fractures than women in the nonosteoporotic control group (P<.001) or than elective osteoporotic women (P = .001) (medians, 5.58, 3.26, and 3.79 pmol/L, respectively; comparisons adjusted for age and estrogen intake). Fifteen patients (50.0%) with hip fractures had deficient vitamin D levels (< or =30.0 nmol/L) and 11 (36.7%) had a parathyroid hormone level greater than 6.84 pmol/L. Levels of N-telopeptide, a marker of bone resorption, were greater in the women with hip fractures than in the elective nonosteoporotic controls (P = .004). CONCLUSIONS: Postmenopausal community-living women who presented with hip fracture showed occult vitamin D deficiency. Repletion of vitamin D and suppression of parathyroid hormone at the time of fracture may reduce future fracture risk and facilitate hip fracture repair. Because vitamin D deficiency is preventable, heightened awareness is necessary to ensure adequate vitamin D nutrition, particularly in northern latitudes.

Aged↗

PubMed1999

Demineralized bone implants for nonunion fractures, bone cysts, and fibrous lesions.

Demineralized bone implants were used to treat eight patients with fracture nonunion, five patients with bone cysts, and eight patients with fibrous lesions. Five of the eight patients with nonunions had had previous unsuccessful attempts at reconstruction. After fixation and implantation with either human or bovine demineralized bone, all eight fractures healed. For those patients with nonunion fractures, mean followup time was 8 years, 5 months, and longest followup was 15 years, 3 months. The cystic lesions in five patients included three typical aneurysmal bone cysts and two recurrent unicameral bone cysts. The aneurysmal bone cysts had excellent healing and bone remodeling. The two unicameral bone cysts were repacked more densely after 1 year and healed. For these patients with bone cysts, the mean followup time was 12 years, 5 months, and longest followup was 15 years, 1 month. The various fibrous lesions in seven of the eight patients were healed within 6 months, with only one requiring repacking. For these patients with fibrous bone lesions, the mean followup time was 9 years, 8 months, and longest followup was 14 years. A biopsy of the lesions in five patients was performed and in two cases showed osteoblasts and new bone around small particles of the implants. These results with long term followup show that allogeneic or xenogeneic demineralized bone implants offer a reasonable alternative for the treatment of typical nonunion fractures, bone cysts, and fibrous lesions of bone.

Adolescent↗

PubMed1999

Medium perfusion enhances osteogenesis by murine osteosarcoma cells in three-dimensional collagen sponges.

In this study, we examined in vitro histogenesis by murine K8 osteosarcoma cells maintained in three-dimensional (3D) collagen sponges. We tested the hypothesis that perfusion of medium enhances cell viability and their biosynthetic activity as assessed by expression of the osteoblastic phenotype and mineral deposition. At intervals, samples were harvested and analyzed histologically, biochemically, and by Northern hybridization for type I collagen, osteopontin (OPN), osteocalcin (OC), and core binding factor alpha 1 (Cbfa1). Histologic evaluation showed greater viability, more alkaline phosphatase (ALP)-positive cells, and more mineralized tissue in the perfused sponges after 21 days. Immunohistological assessment of proliferating cell nuclear antigen revealed 5-fold more proliferating cells in the perfused sponges compared with the controls (p = 0.0201). There was 3-fold more ALP activity in the perfused sponges than the controls at 6 days and 14 days (p = 0.0053). The perfused sponges contained twice the DNA and eight times more calcium than the nonperfused controls after 21 days (p < 0.0001 for both). Northern hybridization analysis revealed more mRNA for collagen type I (2-fold) and 50% more for OC at 14 days and 21 days, whereas OPN and Cbfa1 mRNA expression remained unaffected by the medium perfusion. These results show that medium perfusion had beneficial effects on the proliferation and biosynthetic activity of this osteosarcoma cell line. This system mimics the 3D geometry of bone tissue and has the potential for revealing mechanisms of regulation of osteogenesis.

Alkaline Phosphatase↗

PubMed1999

DHEA and the skeleton (through the ages).

Dehydroepiandrosterone (DHEA) and its sulfate ester, DHEAS, are the most abundant steroids in the human circulation, although their exact biological significance is not completely understood. DHEA(S) levels are high in fetal life, decrease after birth, and show a marked pubertal increase to a maximal level during young adulthood. In healthy adults, DHEAS levels decline to 10-20% of peak levels by age 70 yr. This review summarizes information concerning the role of DHEA in skeletal physiology, including modulation of the skeletal insulin-like growth factor regulatory system, and its effects on secretion of proresorptive cytokines. The pattern of secretion of DHEA throughout the life cycle is discussed, as well as its potential usefulness in specific disease states as an agent with anabolic and antiosteolyic effects on bone.

Adult↗

PubMed1998

Polyethylene glycol/microfibrillar collagen composite as a new resorbable hemostatic bone wax.

Although bone wax is effective at achieving hemostasis, it is nonresorbable, causes a foreign body reaction, and inhibits osteogenesis. We report development of a polyethylene glycol/microfibrillar collagen composite (PEG/MFC) that has inherent hemostatic qualities, is biodegradable, and is compatible with bone repair. PEG/MFC composite (n = 42) was placed in 5 mm cranial defects in New Zealand white rabbits. Hemostasis and healing were compared to unfilled defects (n = 32) and defects filled with standard bone wax (n = 10). Both PEG/MFC and bone wax handled well and stopped bleeding. The polyethylene glycol component was resorbed by 8 h, and the microfibrillar collagen was resorbed over 2 months, eliciting only a minor inflammatory response during the first month. Defects filled with the PEG/MFC composite showed similar amounts of bony regeneration as did unfilled control defects. At 4 weeks, healing bone accounted for 43 +/- 13% in those treated with PEG/MFC and 47 +/- 19% defect area in untreated holes. In contrast, less than 1% of the area was bone in defects filled with bone wax (p < 0.05). PEG/MFC composite provided excellent bony hemostasis and did not inhibit bone growth.

Animals↗

PubMed1998

Angiogenesis in fracture repair.

Fracture of bone disrupts its circulation and leads to necrosis and hypoxia of adjacent bone. Under normal circumstances, fractured bone undergoes the orderly regeneration of its component tissues with complete restoration of mechanical properties. Reestablishment of the circulation is an early event in fracture healing. Several experimental models of protracted, impaired, or compromised healing have been developed to evaluate the effects of angiogenic factors in accelerating or enhancing repair.

Animals↗

PubMed1998

A new in utero sheep model for unilateral coronal craniosynostosis.

Several animal models have been designed in the past to analyze the pathophysiology and management of craniosynostosis, very few of which were intrauterine. Those that were interuterine had problems with either a short gestation or limited availability that prevented most researchers from using them in treatment analysis. We desired to create a biologically sound intrauterine model of craniosynostosis, using an animal with a long gestation and an early calvarial bone formation, which was easy to manipulate in utero, that could be created by any researcher studying this disorder. Using biologic data available regarding growth factors thought to be involved in bone growth and cranial suture closure, we developed a new in utero fetal lamb model for the study of craniosynostosis. Ten 70-day gestation fetal lambs (term gestation 140 days) received a midline coronal incision to expose both coronal sutures. The entire right coronal suture was then excised along with a 4-mm bony margin. In each animal, the site was packed with 25 mg of demineralized sheep bone powder augmented with 50 microg of bone morphogenetic protein-2 (BMP-2) and 1 microg of poly-transforming growth factor-beta. The scalp was closed, and the sheep were returned to the uterus until either 90 or 140 days of gestation. Complete fusion of the right coronal suture occurred in all fetuses by 90 days gestation. In every animal, right-sided frontal bone flattening and supraorbital rim elevation were evident. Histologic analysis showed bony synostosis at the suture site without evidence of suture regeneration. By 140 days, this isolated suture fusion led to marked craniofacial abnormalities including right supraorbital rim elevation, significant frontal bone flattening, a decrease in the anterior-posterior length of the cranial vault, and flattening of the cranial base. In conclusion, we have developed a new model for the study of the secondary effects induced by the process of cranial suture fusion, which produces abnormalities seen in naturally occurring cases of isolated right coronal suture synostosis. In addition, this model confirms that isolated coronal suture fusion alone can lead to the multiple cranial and facial abnormalities seen with this disorder, even in the absence of associated cranial base suture fusions.

Animals↗

PubMed1998

In vitro secretion of cytokines by human bone marrow: effects of age and estrogen status.

It has been proposed that cytokines mediate the acceleration of bone loss following menopause. Because of the intimate relationship between bone marrow stromal cells and bone tissue, it is possible that marrow cells and their products contribute to the bone microenvironment and influence the regulation of bone cell differentiation and activity. We examined the production of cytokines by bone marrow stromal cells from a total of 37 women and 15 men undergoing total hip replacement for noninflammatory joint disease. Low-density mononuclear cells were isolated from bone marrow and were cultured in phenol red-free alpha MEM medium supplemented with 10% FBS and antibiotics. Constitutive secretion of interleukin-6 (IL-6) was positively correlated with age in a series of 8 women and 5 men measured by bioassay (r = 0.98; P < 0.01) and in a series of 18 women and 10 men measured by immunoassay (r = 0.56; P < 0.01). The pattern of cytokine production by bone marrow stromal cells was examined in detail in 23 postmenopausal women, aged 49-88 yr. Basal secretion of immunoreactive IL-6 and IL-11, but not granulocyte-macrophage colony-stimulating factor, increased with time in culture. Exogenous IL-1 beta stimulated secretion of IL-6 and IL-11 in a saturable, dose-dependent manner. Secretion of soluble IL-6 receptor was not correlated with secretion of IL-6, either constitutively or in the presence of IL-1 beta. In 4 of 14 samples, IL-1 beta also stimulated secretion of granulocyte-macrophage colony-stimulating factor. IL-1 beta was undetectable in 7 of 9 cultures during the 2-week culture period. IL-6 did not stimulate secretion of IL-1 beta in the 7 cultures tested. Cells were dependent upon serum for viability and growth and were not sustained by a serum substitute (1% insulin-transferrin-selenium-BSA). Cells grown in medium with 10% FBS and supplemented with 1% insulin-transferrin-selenium-BSA secreted 10-fold more IL-6 than cells grown in serum alone. Marrow from 7 women receiving estrogen replacement therapy showed lower constitutive secretion of IL-6 (75%; P < 0.006) and IL-11 (43%; P < 0.05) than marrow from age-matched controls and had blunted stimulation of IL-6 and IL-11 secretion by exogenous IL-1 beta. These data indicate distinct patterns of cytokine production by human marrow stromal cultures dependent upon age and estrogen status.

Adult↗

PubMed1998

Sex steroids, the insulin-like growth factor regulatory system, and aging: implications for the management of older postmenopausal women.

Aging is associated with profound changes in the growth hormone/insulin-like growth factor (IGF) regulatory system. These include reductions in growth hormone, IGF-I, IGFBP3, and IGFBP-5 and an increase in IGFBP-4. These changes, coupled with rather marked declines in sex steroid production from both the ovary and adrenals may combine to have very deleterious effects on several organ systems in the postmenopausal woman. In particular, the prevalence of two very common diseases, osteoporosis and coronary artery disease, increase dramatically after the cessation of gonadal steroid production. The complex interrelationship between the IGF regulatory system and estrogens/androgens in the postmenopausal period may provide important clues as to the pathophysiology of both these disorders. In this paper, we begin to define the role of IGF-I (and its constituent IGF binding proteins) in skeletal and vascular tissue. Recent experimental data show the effects of estrogen on circulating and tissue IGFs in older individuals. Finally, estrogen replacement therapy affects the IGF regulatory system in postmenopausal women. Although conclusions from early studies remain somewhat preliminary, it is likely that the IGF regulatory system will be a prime target for future studies into the pathogenesis of several age and sex hormone related degenerative disorders.

Aged↗

PubMed1997

Effects of age and estrogen status on the skeletal IGF regulatory system. Studies with human marrow.

Human marrow was obtained as material discarded during total hip replacement and was established in culture with phenol red-free alpha-MEM with 10% fetal bovine serum (FBS) and antibiotics. Insulin-like growth factor I (IGF-I) and its binding proteins were secreted by human marrow cells, in amounts that increased with time in culture. Western ligand blotting showed that insulin-like growth factor binding protein-3 (IGFBP-3) accounted for the majority (approximately 75%) of the secreted binding proteins. Evidence for marrow secretion of BP-3 protease was found by electrophoretic analysis of mixtures of radiolabeled IGFBP-3 and marrow-conditioned media. The amount of constitutive secretion of IGFBP-3 increased with age of the subject (r = 0.97, p = 0.0058). A notable exception was marrow from a postmenopausal women on estrogen replacement therapy (ERT) at the time of surgery; her marrow secreted 89.3 ng/mL after 14 d in vitro, only 38% of the IGFBP-3 that was secreted by cultures from two age-matched peers (208.8 and 285.2 ng/mL). This in vivo effect of estrogen was matched by an in vitro experiment in which 10(-8) M 17-beta estradiol suppressed IGFBP-3 to 60% of the constitutive level. In all cultures of marrow from postmenopausal women, IL-1 beta suppressed IGFBP-3 secretion to either undetectable levels or levels between 11% and 35% of control. Thus, human bone marrow cultures demonstrate components of the skeletal IGF regulatory system: IGF-I, IGF-binding proteins, and evidence of IGFBP-3 proteolysis. These results provide evidence of regulation by both systemic (age, estrogen status) and cytokine (IL-1 beta) factors.

Age Factors↗

PubMed1997

Expression of an extracellular calcium-sensing receptor in human and mouse bone marrow cells.

The cloning of a G protein-coupled, extracellular calcium (Ca2+e)-sensing receptor (CaR) from bovine parathyroid provided direct evidence that Ca2+e-sensing can occur through receptor-mediated activation of G proteins and their associated downstream regulators of cellular function. CaR transcripts and protein are present in various tissues of humans and other mammals that are involved in Ca2+e homeostasis, including parathyroid, kidney, and thyroidal C-cells. The present study was performed to determine whether bone marrow cells express the CaR, since cells within the marrow space could be exposed to substantial changes in Ca2+e related to bone turnover. Using DNA and RNA probes from the human parathyroid CaR cDNA, we identified CaR transcripts of 5.2 and approximately 4.0 kilobases by Northern analysis of poly(A+) RNA from low-density mononuclear cells isolated from whole human bone marrow that are putatively enriched in marrow progenitor cells, including bone cell precursors. In situ hybridization also identified CaR transcripts in the same cell preparations. Reverse transcription-polymerase chain reaction demonstrated > 99% nucleotide identity between transcripts from human bone marrow cells and the corresponding regions of the human CaR cDNA. Antisera specific for several different regions within the extracellular domain of the CaR were reactive with low-density human marrow cells that were either adherent or nonadherent to plastic. About one-third of the adherent, CaR-immunoreactive cells were also positive for alkaline phosphatase, a nonspecific marker of preosteoblasts, osteoblasts, and assorted cells of the colony-forming unit-fibroblast lineage. In addition, a substantial fraction (approximately 60%) of low density murine marrow cells cultured for 1 week at 4.8 mM Ca2+e expressed both CaR immunoreactivity and nonspecific esterase, an enzyme expressed by monocyte/macrophages and fibroblasts. Finally, erythroid precursors and megakaryocytes from murine marrow as well as blood platelets expressed abundant CaR immunoreactivity, while peripheral blood erythrocytes and most polymorphonuclear leukocytes did not. These studies indicate that the CaR is present in low-density mononuclear bone marrow cells as well as in cells of several hematopoietic lineages and could potentially play a role in controlling the function of various cell types within the marrow space.

Animals↗

PubMed1997

Multinucleated cells in pigmented villonodular synovitis and giant cell tumor of tendon sheath express features of osteoclasts.

Pigmented villonodular synovitis (PVNS) and the histologically related lesion giant cell tumor of tendon sheath (GCTTS) are idiopathic, proliferative lesions that can induce osteolysis and formation of bone cysts. These lesions contain two predominant cell types: mononuclear polyhedral cells and multinucleated cells (MNCs). Previous studies demonstrated that the mononuclear cells exhibit phenotypic features consistent with derivation from a monocyte/macrophage lineage. The cell lineage of the MNCs and their relationship to osteoclasts are not known. To characterize the MNCs in these lesions and to establish the relationship of these MNCs to osteoclasts, histological sections from six cases of PVNS and two cases of GCTTS were studied. Mononuclear cells expressed CD14 and HLA-DR, in keeping with their relationship to cells of the monocyte/macrophage lineage. Characterization of the MNCs revealed features associated with an osteoclast phenotype. Seven of the eight specimens contained MNCs that were intensely tartrate-resistant acid phosphatase positive; approximately 5% of the mononuclear cells were tartrate-resistant acid phosphatase positive, and these tended to surround MNCs. MNCs in both lesions reacted strongly with the 23C6 monoclonal antibody that recognizes the alpha V beta 3 integrin (the vitronectin receptor), as did several mononuclear cells surrounding the MNCs. Most MNCs did not express CD14 or HLA-DR. Expression of receptors for calcitonin, a marker for osteoclasts, was detected on MNCs after incubation of sections with 125I-labeled salmon calcitonin and emulsion autoradiography. MNCs in four of six PVNS and two of two GCTTS samples demonstrated specific calcitonin binding. Expression of mRNA for calcitonin receptor was confirmed in all cases by reverse transcriptase polymerase chain reaction. These results demonstrate that MNCs in PVNS and GCTTS express phenotypic features of authentic osteoclasts and suggest that osteoclast-like multinucleated cells can arise in synovial soft tissues remote from bone.

Acid Phosphatase↗

PubMed1996

Chondroinduction of human dermal fibroblasts by demineralized bone in three-dimensional culture.

We designed and tested a three-dimensional device composed of demineralized bone powder (DBP) packed within a bilaminate sponge as an in vitro model for chondroinduction induced by DBP. Control bovine articular chondrocytes deposited cartilage matrix when cultured in these sponges. Human dermal fibroblasts that were seeded onto the composite sponge migrated through the collagen lattice and attached to the particles of DBP. Fibroblasts surrounding the DBP deposited a fine, granular extracellular matrix characterized by metachromatic staining similar to that deposited around the bona fide chondrocytes. Immunohistological analysis showed that the matrix contained chondroitin delta Di-4 sulfate, chondroitin delta Di-6 sulfate, and keratan sulfate epitopes. These components accumulated during the culture period, as quantified by immunochemical analysis of extracted matrix. In contrast, fibroblasts cultured in collagen sponges without DBP did not produce such a cartilage matrix. Control bovine chondrocytes produced these matrix molecules when cultured in the presence or absence of DBP. Thus this composite three-dimensional device facilitated evaluation of chondroinduction in human dermal fibroblasts in vitro.

Animals↗

PubMed1996

Three-dimensional composite of demineralized bone powder and collagen for in vitro analysis of chondroinduction of human dermal fibroblasts.

Implantation of demineralized bone powder (DBP) in muscle or connective tissue stimulates chondrogenesis followed by ectopic bone formation, in this way inducing the differentiation of endochondral bone. A new 3-dimensional in vitro composite sponge was designed to duplicate the packing density of in vivo DBP implants. The composite device, which consists of DBP packed between two layers of a porous collagen lattice, was used to assess the chondroblastic differentiation of human dermal fibroblasts. Important design considerations for this device were biocompatibility, rigidity and ability of cells to penetrate. In this study, collagen concentration and source, irradiation, and lyophilization conditions were varied in fabrication. Human dermal fibroblasts were seeded onto the composite sponge, migrated through the collagen lattice into the packet of DBP, and deposited a metachromatic extracellular matrix amongst the particles of DBP. In contrast, cells cultured in collagen sponges or in composite sponges with inactivated guanidine-extracted DBP did not secrete metachromatic matrix. This new in vitro system will be valuable in defining the mechanism of differentiation by osteoinductive materials and in evaluating the influence of other extracellular components and soluble factors on skeletal differentiation.

Animals↗