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G Lust

Publications and source records attributed to G Lust.

At least 37 records · Page 2Linked to original sources
PubMed1998

Differentiated cellular function in fetal chondrocytes cultured with insulin-like growth factor-I and transforming growth factor-beta.

This study examined fetal chondrocyte proliferation and function following exposure to transforming growth factor-beta and insulin-like growth factor-I. Fetal equine articular chondrocytes of the early third-trimester were isolated and cultured in monolayer conditions, then exposed to 0, 1, 5, or 10 ng/ml transforming growth factor-beta or 0, 10, 50, or 100 ng/ml insulin-like growth factor-I for 48 hours. Proliferative responses were assessed by cell counts and [3H]thymidine uptake into precipitable DNA. Differentiated chondrocyte metabolic activity was determined by sulfated glycosaminoglycan quantitation, 35[SO4] incorporation into precipitable glycosaminoglycan, and proteoglycan molecular sizing by CL-2B column chromatography. Morphological changes seen on phase-contrast microscopy included a larger proportion of rounded cells in monolayer cultures supplemented with insulin-like growth factor-I and cytotoxic changes in cells treated with transforming growth factor-beta. Both insulin-like growth factor-I and transforming growth factor-beta resulted in significant elevations of [3H]thymidine uptake; however, cell numbers did not rise sufficiently over the 48-hour culture period to reach significant levels. Maximum mitogenic responses were evident at 50 and 100 ng/ml insulin-like growth factor-I and 5 ng/ml transforming growth factor-beta. The production of proteoglycan was also enhanced (435%) by exposure to 50 ng/ml insulin-like growth factor-I, and an increased proportion of larger proteoglycan monomer species was evident in cultures treated with 50 and 100 ng/ml insulin-like growth factor-I. A similar dose-response was also evident in cultures treated with transforming growth factor-beta (maximal 164% increase with 5 ng/ml), although the presence of serum in the culture medium altered the pattern of enhanced proteoglycan synthesis to favor the lower concentration of 1 ng/ml (191%). Additionally, larger proteoglycan molecules were synthesized in response to high concentrations of transforming growth factor-beta in serum-free cultures. Significant biochemical changes resulted from the addition of transforming growth factor-beta to fetal chondrocyte cultures; however, monolayer cultures that were treated with transforming growth factor-beta and supplemented with serum began to develop cellular toxicity, including nuclear pyknosis and cytoplasmic fragmentation. Degenerative cellular changes were not evident in cultures treated with insulin-like growth factor-I, and significant differentiated metabolic activity resulted from the presence of insulin-like growth factor-I in the culture medium. These data suggest that the responses of fetal chondrocytes to insulin-like growth factor-I and transforming growth factor-beta were enhanced compared with the responses of chondrocytes derived from postnatal animals and that these metabolically active cells can be primed by endogenous or exogenous growth factors to provide enhanced articular function and repair.

Animals↗
PubMed1998

Expression of the (V+C)- fibronectin isoform is tightly linked to the presence of a cartilaginous matrix.

Fibronectin is encoded by a single gene, but heterogeneity is introduced by alternative splicing of the pre-mRNA. An unique splice variant, designated (V+C)-, which deletes nucleotides encoding the V, III-15 and I-10 segments, has been identified in articular cartilage. In this study, a ribonuclease protection assay was used to quantitate expression of the (V+C)- isoform in eight canine cartilaginous tissues and in chondrocytes cultured as monolayers or in alginate beads. The (V+C)- fibronectin isoform was detected in all cartilaginous tissues examined, ranging from a low of 11% of steady-state fibronectin mRNA in the nucleus pulposus to 71% in the rib. An age dependent increase, from 18% in the epiphyseal cartilage of a newborn to 54% in the articular cartilage of dogs over 10 months of age, was observed. The ubiquitous presence of this isoform in cartilaginous tissues and its absence in all non-cartilaginous tissues examined to date is consistent with a very strong association of the (V+C)- fibronectin isoform with the cartilaginous phenotype. Results from a ribonuclease protection assay using a probe extending into the V region from III-14 were combined with the quantitative information about (V+C)- fibronection expression to develop an over-all profile of splicing within the V region in cartilage. Monolayer culture of articular chondrocytes altered fibronectin splicing patterns. The (V+C)- isoform was rapidly lost and ED-A(+) fibronectin was induced. Three-dimensional culture in alginate beads prevented induction of ED-A(+) fibronection, but failed to sustain expression of the (V+C)- isoform. Thus, some matrix component or structure, lost in cell culture, may be essential to maintain expression of the (V+C)- isoform. The possible relationship of changing patterns of fibronectin isoforms in cultured chondrocytes to maintenance of the differentiated phenotype is discussed.

Alternative Splicing↗
PubMed1998

Two DNA sequences specific for the canine Y chromosome.

Data are presented on the characterization of two nucleotide sequences found exclusively in the DNA of male dogs. In polymerase chain reactions (PCRs) of canine genomic DNA with a decanucleotide primer of arbitrary sequence (OP-W17), two nucleotide segments (650 and 990 bp) were amplified only from male samples, whereas a number of other fragments between 400 and 2500 bp in size were amplified from both male and female samples. The two male-specific segments were cloned and sequenced, and terminal 24mer oligonucleotide primer pairs were synthesized. PCR with these specific primer pairs resulted in amplification of the two male-specific sequences only from DNA samples of 34 male dogs; no product was amplified from 42 samples of females. A segment of the SRY gene previously localized on the Y chromosome could be amplified in DNA samples that had the two new sequences. EcoRI digested genomic male DNA when hybridized with the 650 bp or the 990 bp sequences, resulted in a single band for each on Southern analysis; DNA from females did not yield any bands. Comparisons between the two new sequences and the SRY gene segment revealed no homologies. We concluded that the two new sequences are specific for the canine Y chromosome and do not contain the short characterized segment of the SRY gene.

Animals↗
PubMed1998

Acute synovitis and intra-articular methylprednisolone acetate in ponies.

OBJECTIVE: To determine how acute synovitis, with and without intra-articular methylprednisolone acetate (MPA), affect synthesis of proteoglycan, total protein, and collagen in articular cartilage and total protein synthesis in synovial membrane. DESIGN: Synovitis was induced in 10 ponies by the injection of 0.5 ng lipopolysaccharide (LPS) into the left radiocarpal and midcarpal joints every 2 days for a total of four treatments. Synovitis was documented by clinical examination and synovial fluid analyses. Two days before euthanasia, MPA (0.1 mg/kg) was injected with the last dose of LPS into both the left and right radiocarpal and midcarpal joints of five of these ponies. Proteoglycan synthesis in articular cartilage explants from these joints was measured by incorporation of sodium [35S]sulfate. The size of the proteoglycan monomers and their aggregation with hyaluronan was assessed by size-exclusion chromatography. Protein synthesis in articular cartilage was measured by incorporation of [3H]proline and collagen synthesis by conversion of [3H]proline into [3H]hydroxyproline. Protein synthesis was measured in synovial membrane explants by incorporation of [35S]methionine. RESULTS: Ponies developed carpal effusion and mild lameness accompanied by increased total nucleated cell count and total solids in synovial fluid in response to the LPS injections. Moderate to severe synovial membrane proliferation and inflammation were observed histopathologically in joints injected with LPS but no consistent light-microscopical changes were observed in the articular cartilage from these joints. Intra-articular MPA alone was associated with decreased proteoglycan synthesis and increased protein and collagen synthesis in the cartilage explants. Total protein synthesis by synovial membrane was also increased by MPA alone. In contrast, no differences in protein or proteoglycan synthesis were observed in explants from the joints with synovitis, with or without intra-articular MPA. Treatment with MPA, LPS, and LPS/MPA did not alter proteoglycan aggregate size, but LPS-induced synovitis resulted in an increase in the second largest population of monomers. MPA increased the synthesis of small proteoglycan monomers. CONCLUSION: Based on the methods used, acute synovitis prevented changes induced by intra-articular MPA alone. Results suggested that the effect of intra-articular MPA on joint metabolism was different between inflamed and normal joints. Experimental studies must consider the effect of inflammation, as well as the potential to introduce in vitro culture artifacts when investigating the effect of intra-articular corticosteroids on chondrocyte function.

Acute Disease↗
PubMed1997

An overview of the pathogenesis of canine hip dysplasia.

Canine hip dysplasia (CHD) is an inherited, developmental condition that involves a lack of conformity between the femoral head and acetabulum and invariably leads to osteoarthritis. In this paper, the author briefly reviews what is known about the genetics of CHD, how joint laxity and growth rate are related to development of CHD, and the possibility that CHD is a systemic disease involving multiple joints, not just the hip joint.

Age Factors↗
PubMed1997

Onset of epiphyseal mineralization and growth plate closure in radiographically normal and dysplastic Labrador retrievers.

OBJECTIVE: To determine whether onset of mineralization of the femoral and proximal tibial epiphyses and age at closure of the femoral and acetabular triradiate growth plates was different for Labrador Retrievers that were radiographically normal or that had canine hip dysplasia (CHD). DESIGN: Cohort study. ANIMALS: 36 Labrador Retriever puppies. PROCEDURE: Puppies were radiographed every other day from the time they were 8 to 10 days old until ossification of the femoral heads was apparent. Radiographs were then obtained weekly until puppies were 1 month old and then monthly until puppies were 8 to 12 months old. Age at which mineralization was first observed in the proximal and distal femoral and proximal tibial epiphyses and at which the femoral capital, triradiate acetabular, and distal femoral growth plates were no longer radiographically visible were recorded. Fifteen dogs were euthanatized and necropsied to determine whether dogs had CHD. RESULTS: There were 26 radiographically normal left and right hip joints and 10 dysplastic left and right hip joints. Onset of mineralization of the proximal femoral epiphyses and of the right proximal tibial epiphysis was significantly later in dysplastic than in radiographically normal puppies. The left femoral capital growth plates closed significantly later in dysplastic than in radiographically normal joints, but other differences in growth plate closure were not detected. CLINICAL IMPLICATIONS: Endochondral ossification may be abnormal in dogs with CHD. The disease appears to affect multiple joints, even though it is most evident clinically in the hip joint.

Aging↗
PubMed1997

Variations in composition of cartilage from the shoulder joints of young adult dogs at risk for developing canine hip dysplasia.

OBJECTIVE: To determine whether the composition of cartilage from the shoulder joints of dogs varied with the risk of developing canine hip dysplasia (CHD). DESIGN: Observational study. ANIMALS: 12 skeletally mature (approx 1 year old) Labrador Retrievers. PROCEDURE: Dogs were classified as having a low, moderate, or high risk of developing CHD on the basis of distraction indices. Cartilage was harvested from the craniolateral and weight-bearing regions of the humeral heads, and wet weight per unit area and dry, glycosaminoglycan, and fibronectin contents were determined. RESULTS: Glycosaminoglycan and dry contents did not vary among risk groups. For cartilage from the craniolateral region of the humeral head, wet weight per unit area and fibronectin content increased as risk of developing CHD increased. Wet weight and fibronectin content of cartilage from the weight-bearing region of the humeral head did not vary among risk groups. CLINICAL IMPLICATIONS: Dogs that have a high risk of developing CHD are also more likely to develop osteoarthritis of the shoulder joint. The observed increases in wet weight per unit area and fibronectin content in cartilage from the craniolateral region of the humeral head in dogs at a high risk of developing CHD may be early sings of incipient osteoarthritis.

Animals↗
PubMed1997

Concentrations of keratan sulfate in plasma and synovial fluid from clinically normal horses and horses with joint disease.

OBJECTIVE: To determine whether keratan sulfate concentrations in plasma or synovial fluid from clinically normal horses were different from concentrations in horses with joint disease and whether concentrations varied with type of joint disease. DESIGN: Case-control study. ANIMALS: 67 clinically normal horses, 10 clinically normal foals, and 160 horses with joint disease. PROCEDURE: ELISA was used to measure keratan sulfate concentrations. RESULTS: Mean plasma keratan sulfate concentration (mean +/- SEM, 580 +/- 124 ng/ml) in foals peaked at 10 weeks of age. Mean plasma keratan sulfate concentration in clinically normal horses was 200 ng/ml (95% confidence interval, 157 to 251 ng/ml). Horses with osteochondral (chip) fractures, other closed intraarticular fractures, inflammatory arthritis (synovitis), infectious arthritis, or osteochondrosis had significantly higher plasma keratan sulfate concentrations than did clinically normal horses, but horses with osteoarthritis did not. Breed, gender, and type of joint disease affected keratan sulfate concentration in synovial fluid. Standard-breds with chip fractures of the metacarpophalangeal/ metatarsophalangeal joints had significantly higher keratan sulfate concentrations in synovial fluid than did Thoroughbreds. Keratan sulfate concentrations in synovial fluid from osteoarthritic carpal joints were lower than concentrations in normal carpal joints and tarsocrural joints with inflammatory joint disease. CLINICAL IMPLICATIONS: Keratan sulfate concentration alone was not a specific marker of joint disease but was affected by various joint diseases.

Aging↗
PubMed1997

Five-year longitudinal study on limited food consumption and development of osteoarthritis in coxofemoral joints of dogs.

OBJECTIVE: To examine the effects of limited food intake on frequency and severity of osteoarthritis in coxofemoral joints of labrador Retrievers. DESIGN: Dogs were paired according to gender and body weight, within each litter at 8 weeks of age. One dog of each pair was fed ad libitum. The limit-fed pairmate was fed 75% of the amount eaten the previous day by the ad libitum-fed counterpart. ANIMALS: 48 Labrador Retrievers. PROCEDURE: All dogs received the same diet. Radiographic evaluation of coxofemoral joints for frequency and severity of osteoarthritis were made when dogs were 4 and 6 months and 1, 2, 3, and 5 years old. RESULTS: Radiographic evaluation for osteoarthritis indicated greater frequency and more severity of osteoarthritis in the ad libitum-fed group of dogs. CLINICAL IMPLICATIONS: Analysis of data suggested that limit feeding of dogs over a 5-year period minimizes development of osteoarthritis in the coxofemoral joints.

Animals↗
PubMed1997

Characterization of cartilage metabolic response to static and dynamic stress using a mechanical explant test system.

A new mechanical explant test system was used to study the metabolic response (via proteoglycan biosynthesis) of mature, weight-bearing canine articular cartilage subjected to static and dynamic compressive stresses. Stresses ranging from 0.5 to 24 MPa were applied sinusoidally at 1 Hz for intervals of 2-24 h. The explants were loaded in unconfined compression and compared to age-matched unloaded explants. Both static and dynamic compressive stress significantly decreased proteoglycan biosynthesis (range 25-85%) for all loading time intervals. The inhibition was proportional to the applied stress but was independent of loading time. After rehydration upon load removal, the measured water content of the loaded explants was not different from the unloaded explants for all test variables. Autoradiographic and electron microscopic analysis of loaded explants showed viable chondrocytes throughout the matrix. Our results suggest that the decreased metabolic response of cyclically loaded explants may be dominated by the static component (RMS) of the dynamic load. Furthermore, the observed decreased metabolism may be more representative of the in situ tissue response than that of unloaded explants, in which we found an increasing rate of metabolism for up to 6 days after explant removal.

Animals↗
PubMed1997

Cartilage fibronectin isoforms: in search of functions for a special population of matrix glycoproteins.

Fibronectins are a part of the repertoire of matrix molecules produced by the chondrocyte in order to assemble a functional cartilage matrix. They are encoded by a single gene, but significant protein heterogeneity results from alternative RNA splicing. The population of fibronectin isofroms in adult cartilage is significantly different from fibronectins in other tissues and includes relatively high levels (20-30%) of ED-B(+) fibronectins and high levels (50-80%) of the cartilage specific (V + C)- isoform which lacks the V, III-15 and I-10 segments. Less than 4% of the fibronectins in cartilage are ED-A(+). The synthesis and accumulation of cartilage fibronectins are modulated in response to matrix pathology and to biochemical and mechanical mediators. In addition, alternative splicing patterns are altered when chondrocytes are allowed to dedifferentiate in monolayer culture such that the (V + C)- isoform is lost but the ED-A(+) isoform is reexpressed at high levels. Cartilage fibronectins have the potential to participate in cell signalling via integrin mediated pathways and to interact with other cartilage matrix macromolecules. The tissue-specific splicing pattern gives rise to a unique population of fibronectins within the cartilage. Together, this points to a critical role for cartilage fibronectins in chondrocyte cell biology and the organization of a biomechanically sound matrix. However, the precise function (or functions) of the cartilage fibronectins has (or have) not been defined. This minireview examines current information about the structure, synthesis and interactions of cartilage fibronectins. When possible, potential consequences of the inclusion of the ED-B segment or the exclusion of the V, III-15 and I-10 segments are discussed. The goal is to stimulate critical thought and discussion in the field about cartilage fibronectin isoforms, their function(s) in normal cartilage, and their role(s) in the pathogenesis of cartilage diseases.

Alternative Splicing↗
PubMed1997

Altered biological activity of equine chondrocytes cultured in a three-dimensional fibrin matrix and supplemented with transforming growth factor beta-1.

OBJECTIVE: To determine the effects of transforming growth factor-beta 1 (TGF-beta 1) on the synthesis of DNA, collagen, and proteoglycans (PG) by equine chondrocytes. SAMPLE POPULATION: Articular cartilage obtained from multiple joints of a 4-month-old foal. PROCEDURE: Chondrocytes were isolated by collagenase digestion, cultured in monolayer, trypsinized, and implanted at a cellular density of 10 x 10(6) chondrocytes/ml in a three-dimensional fibrin matrix. Chondrocytes in culture were supplemented with TGF-beta 1 at concentrations of 0, 1, 5, or 10 ng/ml in serum-free medium or medium containing fetal bovine serum (FBS). Total PG accumulation, [35S]-labeled PG synthesis, PG monomer hydrodynamic size, type II collagen production, total DNA content, and [3H]thymidine incorporation into DNA were determined at 7 and 14 days of culture. RESULTS: Chondrocytes maintained a rounded phenotype, dedifferentiating slightly to a more fibroblastic appearance only in medium containing FBS and 10 ng of TGF-beta 1/ml. Type II collagen immunoreaction on day 14 was decreased in the pericellular matrix in cultures containing FBS and 1, 5, and 10 ng of TGF-beta 1/ml, and in all serum-free culture conditions compared to FBS and 0 ng of TGF-beta 1/ml. Total proteoglycan accumulation and [35S]-labeled proteoglycan synthesis in cultures on days 7 and 14 were increased by the addition of exogenous TGF-beta 1 in serum-free conditions and decreased by TGF-beta 1 in FBS-supplemented conditions. Calculation of the partition coefficients for PG indicated that there was synthesis of low molecular weight PG in serum-free conditions and larger sized proteoglycans in FBS-supplemented conditions. Proteoglycan molecular size was unchanged by the addition of TGF-beta 1. Total DNA content of chondrocytes increased with the addition of TGF-beta 1 in FBS-supplemented conditions and decreased in serum-free conditions. CONCLUSIONS: In a solid three-dimensional fibrin matrix, the effects of TGF-beta 1 on chondrocyte biological activity depend on the culture duration and on the presence of FBS in the medium. Stimulatory effects of TGF-beta 1 were most pronounced in serum-free culture conditions with high concentration of TGF-beta 1 (5 and 10 ng/ml) on day 7 and with low concentration of TGF-beta 1 (1 ng/ml) on day 14. CLINICAL RELEVANCE: TGF-beta 1 may not be a suitable growth factor for enhancement of equine articular grafting in sites exposed to serum.

Animals↗
PubMed1996

Fibronectin mRNA splice variant in articular cartilage lacks bases encoding the V, III-15, and I-10 protein segments.

Fibronectin is an extracellular matrix glycoprotein encoded by a single gene. Alternative RNA splicing has been reported at three sites, ED (extra type III domain)-A, ED-B, and the variable or V region. Articular cartilage fibronectin monomers are rarely (ED-A)+, but approximately 25% are (ED-B)+. RNA gel electrophoresis and Northern blot analysis identified two (ED-B)+ and two (ED-B)- fibronectin transcripts in cartilage, each pair differing by approximately 750 bases. This difference results from a previously unreported RNA splicing pattern that eliminates not only the V region but also nucleotides encoding protein segments III-15 and I-10. This new splice variant, which we designate (V+C)-, represents the majority of fibronectin transcripts in equine, canine, and rabbit articular cartilage but is absent in the liver. Reverse transcriptase-polymerase chain reaction analyses of 11 additional equine tissues failed to detect the (V+C)- splice variant, except for very low levels in lymph node, bone, aorta, and skin. Furthermore, chondrocytes grown in monolayer culture maintain high levels of fibronectin expression but stop expressing (V+C)- transcripts over time. The tissue-specific expression pattern of this novel fibronectin isoform suggests that it may have an important function in the matrix organization of cartilage.

Alternative Splicing↗
PubMed1996

Swelling and fibronectin accumulation in articular cartilage explants after cyclical impact.

The objective of this study was to determine if repeated impact could damage living cartilage and lead to osteoarthritis-like changes in its biology. Canine cartilage explants were subjected to impacts of as much as 50 MPa once every 5 seconds for 30 minutes. On each impact cycle, the loading rate was 100 MPa/sec to the assigned peak stress, which was held for 1 second. After impact testing, the cartilage was kept in defined culture for as long as 10 days. Radiosulfate incorporation in the region that received direct impact varied with load 0-4 hours after impact, but it did not vary with load at 20-24 hours after impact. Even so, most explants were visibly damaged by 20 or 50 MPa, and there was subtle evidence of damage from impacts of 5 or 10 MPa. For example, ion-induced swelling in 0.01 M NaCl was increased, suggesting that the physical integrity of the matrix was reduced relative to controls. Self-diffusion of water, measured by proton magnetic resonance imaging was also increased in the deeper zones of the explant, consistent with changes in structure at the molecular level. Ten days after impact, the water content and the fibronectin content of the loaded region of the explant were both increased. In combination, these osteoarthritis-like changes suggested that the physical strength of normal cartilage limits its ability to withstand cyclical impact.

Animals↗
PubMed1996

Effect of methylprednisolone and mechanical loading on canine articular cartilage in explant culture.

The objective of this study was to assess the effect of mechanical load on articular cartilage after in vitro corticosteroid exposure. Canine humeral cartilage was equilibrated for 4 days in defined medium with a serum substitute, then exposed to methylprednisolone sodium succinate for 20 h at 0, 0.01 or 1.0 mg/ml. After a drug-free recovery period, the explants were subjected to 0, 1 or 10 mega pascals (MPa) for 1 out every 5 s for 20 min, then incubated with [35S]-sulfate and [3H]-leucine for 4 h to measure proteoglycan and protein synthesis, respectively. When the loading occurred 22 h after drug exposure, proteoglycan synthesis was inhibited and protein synthesis, was unaffected by the drug. Both were stimulated by load, relative to controls. When the loading was delayed until 142 h after drug exposure, there was no biosynthetic response to load whether or not the explant had been exposed to the drug. Proteoglycan and protein synthesis 142 h after 0 or 0.01 mg/ml were unchanged or slightly higher than at 22 h, in explants which did not receive load. In contrast, biosynthesis were strongly inhibited 142 h after 1.0 mg/ml, and there was a 40% loss of proteoglycan content, relative to 22 h controls. If explants receiving 1.0 mg/ml also received heavy (10 MPa) loads 142 h later, there was a 17% reduction in total dry content suggesting severe matrix damage. These in vitro results suggest that articular load can help maintain normal cartilage metabolism after corticosteroid exposure, but also suggest that heavy loading after a sub-clinical dose can cause a marked loss of matrix solids.

Animals↗
PubMed1996

Use of synthetic canine relaxin to develop a rapid homologous radioimmunoassay.

Canine relaxin (cRlx) was synthesized by a combination of solid-phase methods and sequential site-directed disulfide bond formation. Proof that the intended molecule had been synthesized was obtained by analytical HPLC of the intact and reduced molecule, by amino acid and sequence analysis, and by receptor binding and in vivo mouse interpubic ligament bioassays. Antisera to synthetic cRlx were raised in six male rabbits; these cross-reacted with relaxins of other species, but not with insulin, LH, FSH, hCG, or prolactin (PRL). Three of the antisera neutralized relaxin-induced interpubic ligament formation in estrogen-primed mice in vivo. A new homologous cRlx RIA was developed through the use of rabbit antiserum 79888, synthetic cRlx for standards and 125l-labeled trace, and a goat anti-rabbit lgG-polyethylene glycol precipitant. The new RIA can be completed in 26 h and has a sensitivity of 0.195-0.39 ng cRlx/tube. Intra- and interassay coefficients of variation were 3% and 12.5%. During pregnancy in bitches, serum cRlx rose to about 10 micrograms/ml. Immunoactive cRlx was also detected in serum, colostrum, and milk of lactating bitches, but not in large volumes (100-300 microliters) of serum of pseudopregnant or estrous bitches. Immunoreactive cRlx was also found in seminal plasma, but not in serum, of male dogs. The new homologous cRlx RIA is simple, rapid, sensitive, and specific, and will be used in future studies of canine relaxin physiology.

Amino Acid Sequence↗
PubMed1996

Effect of methylprednisolone acetate on proteoglycan and collagen metabolism of articular cartilage explants.

OBJECTIVE: The effect of different doses of methylprednisolone acetate (MPA) on proteoglycan and collagen metabolism of articular cartilage from normal equine joints was tested in vitro. METHODS: Cultured explants were treated with 0, 0.0004, 0.004, 0.04, 0.4 and 4.0 mg/ml (approximately 10(-6)-10(-2) M) MPA for 72 h. Proteoglycan synthesis was measured by incorporation of sodium [35S]sulfate into proteoglycans and proteoglycan degradation was measured by release of total and radiolabeled proteoglycan into the culture media. The size of the proteoglycans was assessed with size exclusion chromatography under both associative and dissociative conditions. Total protein synthesis was measured by incorporation of [3H]proline and collagen synthesis by formation of [3H]hydroxyproline. RESULTS: Treatments of 0.0004, 0.004, and 0.04 mg/ml MPA caused a significant (p < 0.05) increase in total protein synthesis compared to the control, and doses of 0.0004 and 0.004 mg/ml MPA tended to increase (p < 0.1) collagen synthesis. In contrast, MPA doses of 0.04 mg/ml or less had no effect on proteoglycan synthesis compared to control. Proteoglycan, total protein, and collagen synthesis were severely depressed with the 0.4 and 4.0 mg/ml MPA treatments. The lowest doses of MPA had minimal effect on proteoglycan degradation, while 0.4 and 4.0 mg/ml MPA decreased degradation of total proteoglycan in a dose dependent fashion. Degradation of newly synthesized proteoglycan in the explants was significantly decreased (p < 0.05) by 4.0 mg/ml MPA. There was positive correlation (r = 0.83, p < 0.05) between the effect of MPA on proteoglycan synthesis and the release of total proteoglycan into the culture media. Chromatography under associative conditions showed that treatment with MPA (0.4 and 0.004 mg/ml) decreased the size and increased the polydispersity of aggrecan and induced synthesis of the small nonaggregating proteoglycans. Aggrecan monomers from cartilage treated with MPA included a population of smaller monomers, which resulted in greater polydispersity than those from control cartilage. CONCLUSION: As well as decreasing synthesis, loss of proteoglycan from MPA treated cartilage in vivo may be partly due to the synthesis of a population of smaller proteoglycans than those extracted from untreated cartilage, and these may not interact with hyaluronan and may not be retained permanently in the matrix. Methylprednisolone acetate may affect posttranslational modification of the core protein with the addition of smaller and possibly fewer glycosaminoglycan chains.

Animals↗