text:
Journal of Orthopaedic Research
Published by Wiley
Online ISSN: 1554-527X
Share on Twitter
Quantitative assessment of the effect of 0.05% chlorhexidine on rat articular cartilage metabolismin vitro andIn vivo
AD Reading
Paul Rooney
G. J. S. Taylor
Wound infection remains a problem. Syringe and needle jet lavage of chlorhexidine gluconate 0.05% removed or killed 99.8% of contaminating bacteria within 1 minute in a wound model. In clinical use, however, possible toxicity to articular cartilage is a concern. In an established intact rat patella model in vitro, 1 minute of exposure to chlorhexidine 0.05% and chlorhexidine jet lavage did not significantly alter cartilage metabolism. A 1-hour exposure decreased metabolic activity. In vivo, a 30-minute exposure with or without rinsing produced no impairment of metabolic activity 6 weeks later, suggesting that cartilage has the potential for biological recovery. However, injecting and leaving chlorhexidine 0.05% in the joints was detrimental to the metabolic activity of the articular cartilage as assessed 6 weeks later. Thus, chlorhexidine gluconate 0.05% could be used on normal articular cartilage. Any potential damage from prolonged exposure can be avoided by rinsing after 1 minute.
Read more
Clonal T-cell response against autologous pleomorphic malignant fibrous histiocytoma antigen presented by retrieved HLA-A*0206
Shigeharu Kimura
Yuriko Kozakai
Satoshi Kawaguchi
Toshihiko Yamashita
Towards the goal of identifying tumor-rejection antigens on eradicated tumors in bone and soft tissue sarcomas, we evaluated the immune response against antigens presented by lost HLA class I molecules. Tumor specimens and peripheral blood samples were obtained from a 70-year-old woman with pleomorphic malignant fibrous histiocytoma. Over 1-year culture, a tumor cell line (MFH2004) was established. A B-cell line infected with Epstein-Barr virus (B2004-EBV) was developed from the blood samples. HLA genotypes of B2004-EBVcells were A*0206/2402, B*4006/4601, and C*0102/0801, whereas MFH2004 cells were defective for A*0206, B*4006, and C*0102. Loss of HLA-A2 expression was also proved immunohistochemically in the primary tumor tissues. Lost HLA-A2 in MFH2004 cells was retrieved by transfection of HLA-A*0206 cDNA to develop MFH2004-A2. Attempts to induce CTLs by mixed culture with autologous T cells and MFH2004 cells resulted in failure. In contrast, those with MFH2004-A2 induced CTL clones CTL2004-c6 and CTL2004-c17. These CTL clones specifically killed MFH2004-A2 but not MFH2004 or B2004-EBV in an HLA-A2-restricted manner. These findings suggest that CTL2004-c6 and CTL2004-c17 recognize autologous tumor-rejection antigens presented by HLA-A*0206, which may have been expressed by tumor cells that had been eradicated by the host's immunosurveillance system.
Read more
Author's response to letter to the editor (JOR-13-0436) that refers to "Molecular mechanisms of cell sheet fragment in enhancing bone formation in mandibular distraction"
Dongyang ma
Prevention of ectopic bone formation by local application of ethane-1-hydroxy-1,1-diphosphonate (EHDP): An experimental study in rabbits
L Ahrengart
U Lindgren
This report shows that ectopic bone formation, a serious problem in orthopedic surgery, can be controlled in an animal model by local application of EHDP (disodium-ethane-1-hydroxy-1,1-diphosphonate). The results might be particularly pertinent to the clinical problem of preventing the recurrence of ectopic bone after surgical excision. Male New Zealand white rabbits were treated with immobilization and intermittent passive manipulation of the right knee. The treatment caused bone formation in the quadriceps muscle, which was visible on radiographs after 3 weeks. In this model, the effect of methacrylate implants containing EHDP was studied. A concentration of 16 g EHDP/100 g methacrylate inhibited bone formation in experimental cortical defects. Release of radiolabeled EHDP was studied in an in vitro system. The release of the drug was approximately 20 mg/day and implant initially, decreasing to about 0.1 mg/day/implant after 30 days. Standardized implants containing 16 g EHDP/100 g were then surgically attached to the femur, and the ectopic bone formation created by immobilization and intermittent manipulation was compared with that in rabbits treated with implants but without EHDP. The ectopic bone was measured from lateral and frontal radiograms and from radiograms of serial transverse sections of the thigh. We found that the EHDP implants were capable of preventing major ectopic bone formation in all cases, whereas all rabbits with an implant containing no EHDP had substantial ectopic bone formation at the end of the experiment. There was no difference between groups in the relative amount of cartilage, connective tissue, and normal bone. We conclude that local administration of EHDP may be a useful method for prevention of ectopic bone formation under the conditions and time employed.
Read more
T2 relaxation time and delayed gadolinium-enhanced MRI of cartilage (dGEMRIC) of human patellar cartilage at 1.5 T and 9.4 T: Relationships with tissue mechanical properties
Eveliina Lammentausta
P Kiviranta
Mikko J Nissi
Jukka Jurvelin
Quantitative magnetic resonance imaging (MRI) techniques have been developed for noninvasive assessment of the structure of articular cartilage. T2 relaxation time is sensitive to the integrity and orientation of the collagen network, while T1 relaxation time in presence of Gd-DTPA2- (dGEMRIC) reflects the proteoglycan content of cartilage. In the present study, human patellar cartilage samples were investigated in vitro to determine the ability of MRI parameters to reveal topographical variations and to predict mechanical properties of cartilage at two different field strengths. T2 and dGEMRIC measurements at 1.5 T and 9.4 T were correlated with the static and dynamic compressive moduli at six anatomical locations of the patellar surface. Statistically significant linear correlations were observed between MRI and mechanical parameters at both field strengths, especially between T2 and Young's modulus. No significant difference was found between the T2 measurements at different field strengths in predicting mechanical properties of the tissue. Topographical variation of T2 values at both field strengths was similar to that of Young's moduli. The current results demonstrate the feasibility of quantitative MRI, particularly T2 mapping, to reflect the mechanical properties of human patellar cartilage at both field strengths.
Read more
Analysis of a retrieved UHMWPE acetabular cup crosslinked in air with 1000 kGy of gamma radiation
Nobuhiko Sugano
M Saito
T Yamamoto
Aiguo Wang
The oxidation and degradation that accompany high dose gamma irradiation in air for crosslinking and sterilization are important because they could affect the clinical performance of polyethylene total joint implants. We report a clinical case of a 1000 kGy gamma-irradiated, highly crosslinked ultra-high molecular weight polyethylene acetabular cup retrieved 24 years postoperatively. Performance evaluations included absorbed radiation dose, total wear penetration, average wear rate, crystallinity, oxidation, and equibiaxial tensile properties. A retrieved acetabular cup of the same grade of polyethylene but gamma-sterilized using a conventional dose was used as control. The highly crosslinked and control cups took in about 1300 and 30 kGy of radiation, respectively, as measured using a trans-vinylene index. Direct dimensional measurements revealed average wear rates of the highly crosslinked and control cups were 0.04 and 0.06 mm/year, respectively. The oxidation index of the highly crosslinked cup was very high (0.679), but equivalent to that of a 1000 kGy irradiated reference cup. The retrieved highly crosslinked cup showed much higher equibiaxial ultimate tensile strength than the retrieved control cup. Based on these observations, the increased wear resistance and equibiaxial tensile properties that resulted from extensive crosslinking in the presence of air were partially offset by the adverse effects caused by immediate oxidation during the process.
Read more
"Shear movement at the fracture site delays healing in a diaphyseal fracture model" by Peter Augat, Johannes Buger, Sandra Schorlemmer, Thomas Henke, Manfred Peraus, Lutz Claes [J Orthop Res 2003;21:1011–17]
S.-H. Park
Re: Effect of Quercetin on Bone Formation: JOR 25:1061-1066, 2008 and Other Publications Reporting the Effects of Various Materials on Bone Formation in Rabbit Calvarial Defects
Ricky Wong
High concentration and bioactivity of vancomycin and aztreonam eluted from Simplex™ cement spacers in two-stage revision of infected hip implants: A study of 46 patients at an average follow-up of 107 days
Pang-Hsin Hsieh
Yu-Han Chang
Shih-Hao Chen
Chun-Hsiung Shih
This study investigated the release of antibiotics in vivo, from an articulating polymethylmethacrylate (PMMA) spacer used in two-stage revision arthroplasty of infected hip implants. Forty-six patients who underwent two-stage revision hip arthroplasty for infections were managed with an interim PMMA spacer loaded with a high dose of vancomycin and aztreonam. Serum and aliquots of drainage collected after the first-stage surgery, and joint fluid obtained at the time of the second-stage surgery were analyzed for antibiotic concentrations by high performance liquid chromatography and bioactivity by tube dilution bioassay. Following implantation, the highest levels of antibiotics were measured in aliquots of drainage on the first day (vancomycin: 1538.0 +/- 243.6 microg/mL; aztreonam: 1003.5 +/- 323.5 microg/mL), decreasing to 571.9 +/- 169.4 microg/mL for vancomycin and 313.6 +/- 88.3 microg/mL for aztreonam after 7 days. Antibiotic concentrations in serum were very low (vancomycin: 0.58 +/- 0.2 microg/mL, range: 0.1-1.6 microg/mL; aztreonam: 0.46 +/- 0.3 microg/mL, range: 0.1-0.9 microg/mL at 24 h) and there was no systemic adverse effect. At a mean 107 days after the first-stage surgery, the concentrations of antibiotics in joint fluid were well above the minimal inhibitory concentration of most common microorganisms. The released antibiotics were bioactive against the test organisms. Based on the observed results, we confirmed the safety and effectiveness of in vivo drug delivery from antibiotic-impregnated PMMA hip spacers.
Read more
In vivo release of the antimicrobial peptide hLF1-11 from calcium phosphate cement
Hein P Stallmann
Ronald de Roo
Chris Faber
Paul I.J.M. Wuisman
We studied the release of human lactoferrin 1-11 (hLF1-11), a potent antimicrobial peptide, in an animal model. Calcium phosphate cement with 50 mg/g hLF1-11 was injected into the femoral canal of 12 rabbits. One, 3, and 7 days later, four animals were terminated, and the femora excised. Sections of bone and cement were removed for histological analysis. We used liquid chromatography-mass spectrometry/mass spectrometry for semiquantitative determination of the hLF1-11 concentration. Blood samples were drawn for leukocyte count and differentiation to identify a potential immunomodulating effect of hLF1-11. After an initial burst release, the hLF1-11 concentration in cement and bone decreased steadily. This in vivo release profile is consistent with earlier in vitro studies. Tissue ingrowth into the cement, without signs of inflammation or necrosis, was observed. Leukocytosis or a shift in leukocyte differentiation did not occur. The carrier released over 99% of the hLF1-11, resulting in peak concentrations at the cement-bone interface. This indicates that hLF1-11 could become a valuable prophylactic agent in osteomyelitis treatment.
Read more
Measurement of blood flow to the tibial diaphysis using 11-?m radioactive microspheres. A comparative study in the adult rabbit
Paul D. Triffitt
Paul J Gregg
The commonly used size of microsphere for bone blood flow estimation is 15 microns, because it has appeared to be the smallest size that is not subject to significant nonentrapment in bone. Soft-tissue studies suggest that it is microspheres of 9-10 microns or less that pass through peripheral tissues and give low calculated flows, whereas many vessels passing into and within cortical bone are less than 15 microns in diameter. We have therefore performed a comparison between microspheres approximately 15 and 11 microns in average size. Blood flows to the cortex and marrow of the tibial diaphysis, and to the skeletal muscle of the anterior compartment, were obtained in six adult New Zealand White rabbits by the reference sample technique, injecting microspheres of 16.5 +/- 0.1 microns and 11.3 +/- 0.1 microns simultaneously. The calculated cortical flows averaged 2.07 and 2.51 ml/min/100 g, respectively, and the marrow flows 26.63 and 24.92 ml/min/100 g. Mean skeletal muscle flows were 15.57 and 14.54 ml/min/100 g, respectively. There were no significant differences between the calculated flows for the two sizes of microsphere. Thus, the smaller microspheres do not appear by this method to be subject to significant nonentrapment, and they are therefore suitable for blood flow measurement in these tissues.
Read more
Indium-111 leukocyte scintigraphic detection of subclinical osteomyelitis complicating delayed and nonunion long bone fractures: A prospective study
John L. Esterhai
Stephen R. Goll
Kevin E. McCarthy
R. Bruce Heppenstall
Twenty patients were studied prospectively with indium-labeled leukocyte imaging to evaluate its effectiveness in differentiating noninfected delayed or nonunion from osteomyelitis complicating these entities. All patients underwent an open surgical procedure within 24 h of the scan. Bone specimens from the nonunion site were obtained for microbiological and histological analysis to confirm the presence or absence of osteomyelitis. In these twenty patients, the sensitivity of the indium scintigraphy was 100%, the specificity 100%, and the overall accuracy 100%. Indium-labeled leukocyte scintigraphy is significantly more accurate than 99mtechnetium and 67gallium imaging had been, when studied earlier, in detecting subclinical osteomyelitis complicating nonunion. Indium-labeled leukocyte scintigraphy should supplant sequential technetium and gallium studies in this patient population when the surgeon must determine whether subclinical osteomyelitis is complicating fracture management of delayed and nonunions.
Read more
Schipplein OD, Andriacchi TP. Interaction between active and passive knee stabilizers during level walking. J Orthop Res.9: 113-119
O.D. Schipplein
Thomas P Andriacchi
The gait of normal subjects and patients with varus deformities at the knee was studied by analyzing the interaction between the dynamic (muscular) and passive (ligamentous) restraints affecting lateral stability of the knee. A statistically determinant model predicted that the midstance-phase adducting moment during normal gait would cause lateral knee joint opening if either antagonistic muscle force and/or pretension in the lateral soft tissues were not present at the knee. The patient group tended to compensate for a high midstance-phase adducting moment by walking with a style of gait that demanded more muscle force (greater flexion-extension moments). This walking style reduced the chance of lateral joint opening. It can be speculated that this style of gait would help to maintain equilibrium at the knee. The higher muscle force would aid in resisting the adducting moment, keeping the joint closed laterally and thus increasing the stability of the knee.
Read more
Vascular endothelial growth factor 121 and 165 in the subacromial bursa are involved in shoulder joint contracture in type II diabetics with rotator cuff disease
Akiyoshi Handa
Masafumi Gotoh
Kazutoshi Hamada
Hiroaki Fukuda
Vascular endothelial growth factor (VEGF) is a glycoprotein that plays an important role in neovascularization and increases vascular permeability. We reported that VEGF is involved in motion pain of patients with rotator cuff disease by causing synovial proliferation in the subacromial bursa (SAB). The present study investigates whether VEGF is also involved in the development of shoulder contracture in diabetics with rotator cuff disease. We examined 67 patients with rotator cuff disease, including 36 with complete cuff tears, 20 with incomplete tears, and 11 without apparent tears (subacromial bursitis). The patients were into groups according to the presence or absence of diabetes (14 type II diabetics and 53 non-diabetics). Specimens of the synovium of the SAB were obtained from all patients during surgery. Expression of the VEGF gene in the synovium of the subacromial bursa was evaluated by using the reverse transcriptase polymerase chain reaction. The VEGF protein was localized by immunohistochemistry, and the number of vessels was evaluated based on CD34 immunoreactivity. The results showed that VEGF mRNA was expressed in significantly more diabetics (100%, 14/14) than in non-diabetics (70%, 37/53) (P=0.0159, Fisher's test). Investigation of VEGF isoform expression revealed VEGF121 in all 14 diabetics and in 37 of the 53 non-diabetics, VEGF165 in 12 of the 14 diabetics and in 21 of the 53 non-diabetics, and VEGF189 in 1 of the 14 diabetics and in 2 of the 53 non-diabetics. No VEGF206 was expressed in either group. VEGF protein was localized in both vascular endothelial cells and synovial lining cells. The mean number of VEGF-positive vessels and the vessel area were also significantly greater in the diabetics (p<0.015, Mann-Whitney U test). Synovial proliferation and shoulder joint contracture were more common in the diabetics (P=0.0329 and P=0.073, respectively; Fisher's test). The mean preoperative range of shoulder motion significantly differed in terms of elevation between two groups: 103.8 degrees in diabetics and 124.9 degrees in no diabetics (p=0.0039 Mann-Whitney U test). In contrast, external rotation did not significantly differ: 44 degrees in diabetics and 49 degrees in non-diabetics (p=0.4957, Mann-Whitney U test). These results suggest that VEGF121 and VEGF165 expression in the SAB is responsible for the development of shoulder joint contracture, especially in elevation, among type II diabetic patients with rotator cuff disease.
Read more
Retention of I-125-labeled recombinant human bone morphogenetic protein-2 by biphasic calcium phosphate or a composite sponge in a rabbit posterolateral spine arthrodesis model
John Louis-Ugbo
Hak-Sun Kim
Scott Boden
Steve Peckham
The purpose of this study was to characterize the retention kinetics of recombinant human bone morphogenetic protein-2 (rhBMP-2) applied to two calcium-based delivery matrices. Biphasic calcium phosphate (BCP) and a composite containing BCP in an absorbable collagen sponge (BCP/ACS) were evaluated using a spinal fusion model in rabbits. rhBMP-2 labeled with radioactive iodine (¹²⁵I) was used as a tracer to assess in vivo retention of rhBMP-2 in the presence of these materials (nine animals per material studied). Over a 36 day study period, animals were assessed for the following: percent administered dose retained at the implant site as measured by scintigraphic imaging (counting) with a gamma camera (all animals), radiography of the implant site (all animals), radioactivity in blood and plasma (all animals), and radioactivity in the urine and feces (three animals for each material). Radioactivity data were corrected for the decay of ¹²⁵I and the attenuation between the implant in vivo and the gamma camera.
Read more
A Novel Locus for Adolescent Idiopathic Scoliosis on Chromosome 12p
Cathleen L Raggio
Philip F Giampietro
Seth Dobrin
Robert Blank
Adolescent idiopathic scoliosis (AIS) is a common disorder with strong evidence for genetic predisposition. Quantitative trait loci (QTLs) for AIS susceptibility have been identified on chromosomes. We performed a genome-wide genetic linkage scan in seven multiplex families using 400 marker loci with a mean spacing of 8.6 cM. We used Genehunter Plus to generate linkage statistics, expressed as homogeneity (HLOD) scores, under dominant and recessive genetic models. We found a significant linkage signal on chromosome 12p, whose support interval extends from near 12 pter, spanning approximately 10 million bases or 31 cM. Fine mapping within the region using 20 additional markers reveals maximum HLOD = 3.7 at 5 cM under a dominant inheritance model, and a split peak maximum HLOD = 3.2 at 8 and 18 cM under a recessive inheritance model. The linkage support interval contains 95 known genes. We found evidence suggestive of linkage on chromosomes 1, 6, 7, 8, and 14. This study is the first to find evidence of an AIS susceptibility locus on chromosome 12. Detection of AIS susceptibility QTLs on multiple chromosomes in this and other studies demonstrate that the condition is genetically heterogeneous.
Read more
Hyaluronan Inhibits Matrix Metalloproteinase-13 in Human Arthritic Chondrocytes via CD44 and P38
Sohel M Julovi
Hiromu Ito
Kohei Nishitani
Takashi Nakamura
We investigated the effects of hyaluronan (HA) on interleukin-1β (IL-1β)-stimulated matrix metalloproteinase (MMP)-13 production in human chondrocytes from patients with osteoarthritis (OA) or rheumatoid arthritis (RA). Secreted levels of MMP-13 in conditioned media were detected by immunoblotting, while intracellular MMP-13 synthesis in articular cartilage was evaluated by immunofluorescence microscopic analysis. Mitogen-activated protein kinases (MAPKs), p38, extracellular signal-regulated kinases (ERK), and c-jun NH2-terminal kinase (JNK) were assessed by Western blotting. IL-1β (2 ng/ml) stimulates the secretion of MMP-13 in both OA and RA chondrocytes. Inhibition studies using specific MAPK inhibitors revealed that IL-1β induced MMP-13 via p38 in both OA and RA chondrocytes. HA down-regulates IL-1β-stimulated MMP-13 and phosphorylated p38 (p-p38) in a dose-dependent manner (0.1, 1, 2, and 4 mg/ml). When used at 4 mg/ml, HA inhibits p-p38 phosphorylation by more than 60%. In response to IL-1β, RA chondrocytes express a higher level of p-p38 than that of OA chondrocytes. Inhibition of CD44, using a blocking antibody, significantly reversed the inhibitory effect of HA on both MMP-13 and p-p38. Our study clearly shows that HA inhibits IL-1β-induced MMP-13 via its principal receptor, CD44, and subsequent intracellular p38 MAPK signaling in OA and RA chondrocytes.
Read more
Interface membrane fibroblasts around aseptically loosened endoprostheses express MMP-13
Susanne Mayer-Wagner
Hans Gollwitzer
Dirk Wernicke
Willy Hofstetter
The objective of this article was to assess whether matrix metalloproteinase-13 (MMP-13) is produced by cells of the peri-implant interface tissues and to further characterize these cells. Tissue specimens were collected from the bone-prosthesis interface at the time of revision surgery of clinically loosened hip and knee arthroplasties (n = 27). Synovial tissues from osteoarthritic patients and young patients with mild joint deformity were used as controls (n = 6). Tissue samples were fixed in 4% PFA, decalcified with EDTA, and embedded in paraffin. Sections (4 microm) were stained with hematoxylin/eosin and for the osteoclastic marker enzyme tartrate resistant acid phosphatase. Monocytes/macrophages were characterized with a monoclonal antibody against CD68 and mRNAs encoding MMP-13 and alpha(1) collagen I (COL1A1) were detected by in situ hybridization. Cells expressing transcripts encoding MMP-13 were found in 70% of the interface tissues. These cells colocalized with a cell population expressing COL1A1 mRNA, and were fibroblastic in appearance. MMP-13 expressing cells were found in the close vicinity of osteoclasts and multinuclear giant cells. No signals for transcripts encoding MMP-13 were detected in multinuclear giant cells or in osteoclasts. Control tissues were negative for transcripts encoding MMP-13 mRNA. Fibroblasts of the interface from aseptically loosened endoprostheses selectively express MMP-13. By the expression and the release of MMP-13, these fibroblastic cells may contribute to the local degradation of the extracellular matrix and to bone resorption.
Read more
Reduced cortical bone mass in mice with inactivation of interleukin-4 and interleukin-13
Carl-Johan Silfverswärd
Sune Larsson
Claes Ohlsson
Olle Nilsson
The aim of the present study was to study the in vivo role of IL-4 and IL-13 on bone metabolism. The skeletal phenotypes of male and female IL-13(-/-) (n = 7+7), IL-4(-/-)IL-13(-/-) (n = 7+7), and WT (n = 7+7) mice were compared. Analysis was made at 6 weeks of age (juvenile) by pQCT, and at 20 weeks of age (adult) by pQCT, biomechanical testing, and by S-IGF-1 and S-Osteocalcin measurements. The skeletal phenotype was affected only in adult male IL-4(-/-)IL-13(-/-) mice. These animals displayed a reduction in cortical bone mineral content (BMC) of both the tibia and the femur, as measured by mid-diaphyseal pQCT scans, compared with WT mice (tibia -8.2%; femur -8.5%; p < 0.01). This reduction in cortical BMC was due to a decreased cross-sectional area as a result of a reduced cortical thickness. The mechanical strength of the cortical bone, tested by three-point-bending at the mid-diaphyseal region of the femurs, demonstrated a significant reduction of displacement at failure (-11.4%), maximal load at failure (-10.6%), and total energy until failure (-29.4%). S-IGF-1 and S-Osteocalcin levels as well as trabecular bone mineral density (tvBMD) were unaffected in adult male IL-4(-/-)IL-13(-/-) mice. IL-4(-/-)IL-13(-/-) male mice show adult onset reduction of cortical bone mass and strength, indicating that the two anti-inflammatory Th(2) cytokines IL-4 and IL-13 are involved in the regulation of bone remodeling.
Read more
Ibuprofen Upregulates Expressions of Matrix Metalloproteinase-1,-8,-9, and-13 without Affecting Expressions of Types I and III Collagen in Tendon Cells
Wen-Chung Tsai
Chih-Chin Hsu
Hsiang-Ning Chang
Jong-Hwei Pang
Nonsteroidal antiinflammatory drugs are widely used to treat sports-related tendon injuries or tendinopathy. This study was designed to investigate the effect of ibuprofen on expressions of types I and III collagen, as well as collagen-degrading enzymes including matrix metalloproteinase (MMP)-1, -2, -8, -9, and -13. Rat Achilles tendon cells were treated with ibuprofen and then underwent MTT [3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide] assay. Reverse transcription-polymerase chain reaction was used to evaluate mRNA expressions of types I and III collagen, MMP-1, -2, -8, -9, and -13. Protein expressions of types I and III collagen, MMP-1, -8, and -13 were determined by Western blot analysis. Gelatin zymography was used to evaluate the enzymatic activities of MMP-2 and MMP-9. The results revealed that ibuprofen upregulated expressions of MMP-1, -8, -9, and -13, both at mRNA and protein levels. There was no effect of ibuprofen on mRNA and protein expressions of types I and III collagen. Gelatin zymography revealed that the enzymatic activity of MMP-9 was upregulated after ibuprofen treatment. In conclusion, ibuprofen upregulates the expressions of collagenases including MMP-1, -8, -9, and -13 without affecting the expressions of types I and III collagen. These findings suggest a molecular mechanism potentially accounting for the inhibition of tendon healing by ibuprofen.
Read more
Intra-Articular Injection of Hyaluronan Restores the Aberrant Expression of Matrix Metalloproteinase-13 in Osteoarthritic Subchondral Bone
Nobuyuki Hiraoka
Kenji A Takahashi
Yuji Arai
Toshikazu Kubo
Subchondral bone is a candidate for treatment of osteoarthritis (OA). We investigated the effects of intra-articular injection of hyaluronan (IAI-HA) on subchondral bone in rabbit OA model. OA was induced by anterior cruciate ligament transection, with some rabbits receiving IAI-HA. OA was graded morphologically, and expression of mRNA was assessed by real-time RT-PCR. Tissue sections were stained with hyaluronan-binding protein, and penetration of fluorescent hyaluronan was assessed. The in vitro inhibitory effect of hyaluronan on MMP-13 was analyzed in human osteoarthritic subchondral bone osteoblasts (OA Ob) by real-time RT-PCR and ELISA. Binding of hyaluronan to OA Ob via CD44 was assessed by immunofluorescence cytochemistry. Expression of MMP-13 and IL-6 mRNA in cartilage and subchondral bone, and morphological OA grade, increased over time. IAI-HA ameliorated the OA grade and selectively suppressed MMP-13 mRNA in subchondral bone. IAI-HA enhanced the hyaluronan staining of subchondral bone marrow cells and osteocyte lacunae. Fluorescence was observed in the subchondral bone marrow space. In OA Ob, hyaluronan reduced the expression and production of MMP-13, and anti-CD44 antibody blocked hyaluronan binding to OA Ob. These findings indicate that regulation of MMP-13 in subchondral bone may be a critical mechanism during IAI-HA.
Read more
Autologous protein solution inhibits MMP-13 production by IL-1β and TNFα-stimulated human articular chondrocytes
Jennifer Woodell-May
Andrea Matuska
Megan Oyster
Jacy Hoeppner
Catabolic inflammatory cytokines are prevalent in osteoarthritis (OA). The purpose of this study was to evaluate an autologous protein solution (APS) as a potential chondroprotective agent for OA therapy. APS was prepared from platelet-rich plasma (PRP). The APS solution contained both anabolic (bFGF, TGF-β1, TGF-β2, EGF, IGF-1, PDGF-AB, PDGF-BB, and VEGF) and anti-inflammatory (IL-1ra, sTNF-RI, sTNF-RII, IL-4, IL-10, IL-13, and IFNγ) cytokines but low concentrations of catabolic cytokines (IL-1α, IL-1β, TNFα, IL-6, IL-8, IL-17, and IL-18). Human articular chondrocytes were pre-incubated with the antagonists IL-1ra, sTNF-RI, or APS prior to the addition of recombinant human IL-1β or TNFα. Following exposure to inflammatory cytokines, the levels of MMP-13 in the culture medium were evaluated by ELISA. MMP-13 production stimulated in chondrocytes by IL-1β or TNFα was reduced by rhIL-1ra and sTNF-RI to near basal levels. APS was also capable of inhibiting the production of MMP-13 induced by both IL-1β and TNFα. The combination of anabolic and anti-inflammatory cytokines in the APS created from PRP may render this formulation to be a potential candidate for the treatment of inflammation in patients at early stages of OA.
Read more
IL-1 beta induces COX2, MMP-1, -3 and -13, ADAMTS-4, IL-1 beta and IL-6 in human tendon cells
M Tsuzaki
G Guyton
William Garrett
Albert J Banes
Overuse injuries and trauma in tendon often involve acute or chronic pain and eventual matrix destruction. Anti-inflammatory drugs have been used as a treatment, however, the cellular and molecular mechanisms of the destructive processes in tendon are not clearly understood. It is thought that an inflammatory event may be involved as an initiating factor. Mediators of the inflammatory response include cytokines released from macrophages and monocytes. Interleukin-1 beta (IL-1 beta) is a candidate proinflammatory cytokine that is active in connective tissues such as bone and cartilage. We hypothesized that tendon cells would express receptors and respond to IL-1 beta in an initial "molecular inflammation" cascade, that is, connective tissue cell expression of cytokines that induce matrix destructive enzymes. This cascade results in expression of matrix metalloproteinases (MMPs) and aggrecanases that may lead to matrix destruction. Normal human tendon cells from six patients were isolated, grown to quiescence and treated with human recombinant IL-1 beta in serum-free medium for 16 h. Total RNA was isolated and mRNA expression assessed by semiquantitative RT-PCR. IL-1 beta (1 nM) induced mRNAs for cyclooxygenase 2 (COX2), MMP-1, -3, -13 and aggrecanase-1 as well as IL-1 beta and IL-6, whereas mRNAs for COX1 and MMP-2 were expressed constitutively. The IL-1 beta-treated tendon cells released prostaglandin E(2) (PGE(2)) in the medium, suggesting that the inducible COX2 catalyzed this synthesis. Induction of PGE(2) was detectable at 10 pM IL-1 beta. IL-1 beta also stimulated MMP-1 and -3 protein secretion. Induction of MMP-1 and -3 was detectable at 10 pM IL-1 beta. Post-injury or after some other inciting events, exogenous IL-1 beta released upon bleeding or as leakage of local capillaries may drive a proinflammatory response at the connective tissue cell level. The resulting induction of COX2, MMP-1 and -3 may underscore a potential for nonlymphocyte-mediated cytokine production of MMPs that causes matrix destruction and a loss of tendon biomechanical properties. Endogenous IL-1 beta might contribute to the process through a positive feedback loop by stimulating expression and accumulation of MMPs in the tendon matrix.
Read more
Synoviocytes Are More Sensitive Than Cartilage to the Effects of Minocycline and Doxycycline on IL-1 alpha and MMP-13-Induced Catabolic Gene Responses
Lisa A Fortier
Tatiana Motta
Robert A Greenwald
Karen G Mayr
The objective of this study was to determine the primary articular tissue target of doxycycline and minocycline. Synoviocytes-cartilage cocultures (n = 4) were treated with MMP-13 (25 ng/mL medium) or IL-1 (1.0 ng/mL medium) for 24 h. Doxycycline (4.3, 0.43, 0.043 microM) or minocycline (10, 1.0 or 0.1 microM) were then added and cultures were continued for 96 h. Cartilage and media were analyzed for GAG content. Quantitative PCR was used to measure cartilage MMP-3, MMP-13, aggrecan, COL2A1, ADAMTS-4, and ADAMTS-5 expression, and synoviocyte MMP-3, MMP-13, ADAMTS-4, and ADMATS-5 expression. Total and active MMP-3, MMP-13, and ADAMTS 4/5 enzymes were measured in culture medium. All concentrations of doxycycline and minocycline diminished GAG accumulation in the media. All concentrations of minocycline, but only the highest concentration of doxycycline decreased MMP-3 and MMP-13 expression in synoviocytes but not cartilage, and basal ADAMTS-5 mRNA levels in both synoviocytes and cartilage. Only minocycline decreased active MMP-13 protein in synoviocytes. In summary, the protective effects of tetracycline compounds are more pronounced in synoviocytes than cartilage, and following minocycline compared to doxycycline. Studies to determine the molecular mechanism of action of the tetracyclines in synoviocytes might lead to the design of targeted therapeutics for the treatment of OA or RA.
Read more
Interleukin-4 and interleukin-13 potentiate interleukin-1 induced secretion of interleukin-6 in human osteoblast-like cells
Carl-Johan Silfverswärd
Anders Frost
Helena Brändström
Osten Ljunggren
Formation of interleukin-6 (IL-6) in osteoblasts and bone marrow stromal cells is believed to regulate osteoclast recruitment. The anti-inflammatory cytokines interleukin-4 and -13 (IL-4 and IL-13) stimulate IL-6 production in human osteoblasts. We investigated the relative potencies, and synergistic effects, between IL-4, IL-13 and interleukin-1 (IL-1) on IL-6 formation in human osteoblast-like cells. Isolated human osteoblast-like cells were incubated for 72 h in the presence of various concentrations of IL-4, IL-13 and IL-1, and IL-6 secretion was measured by ELISA. All cytokines stimulated the secretion of IL-6. The rank order of potency was IL-1>IL-4>IL-13. There were no additive or synergistic effects between IL-4 and IL-13. However, co-stimulation with IL-1 and IL-4 resulted in a marked synergistic effect on IL-6 secretion. Co- stimulation with IL-1 and IL-13 gave a minor synergistic effect. In conclusion, IL-4/13 synergistically potentiates IL-1 induced secretion of IL-6 in human osteoblast-like cells.
Read more
Unfavorable Hip Stress Distribution After Legg-Calve-Perthes Syndrome: A 25-Year Follow-Up of 135 Hips
Boštjan Kocjančič
Andrej Moličnik
Vane Antolic
Rok Vengust
To study the effect of hip and pelvis geometry on development of the hip after Perthes disease, we determined the resultant hip force and contact hip stress distribution in a population of 135 adult hips of patients who had been treated for Perthes disease in childhood. Contra-lateral hips with no record of disease were taken as the control population. Biomechanical parameters were determined by mathematical models for resultant hip force in one-legged stance and for contact hip stress, which use as an input the geometrical parameters assessed from anteroposterior radiographs. The mathematical model for stress was upgraded to account for the deviation of the femoral head shape from spherical. No differences were found in resultant hip force and in peak contact hip stress between the hips that were in childhood subject to Perthes disease and the control population, but a considerable (148%) and significant (p < 0.001) difference was found in the contact hip stress gradient index, expressing an unfavorable, steep decrease of contact stress at the lateral acetabular rim. This finding indicates an increased risk of early coxarthritis in hips subject to Perthes disease. © 2013 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res.
Read more
Single-nucleotide polymorphism in Turkish patients with adolescent idiopathic scoliosis: Curve progression is not related with MATN-1, LCT C/T-13910, and VDR BsmI
Hurriyet Yılmaz
Coskun Zateri
Ahmet Uludag
Ozturk Ozdemir
The role of genetics in the etiopathogenesis of adolescent idiopathic scoliosis (AIS) is unclear. In this study, we investigated the relationship between AIS and polymorphisms in MATN-1, LCT C/T-13910, and VDR BsmI genes. 53 Turkish adolescents with diagnosed AIS and 54 healthy adult individuals were included in the study. MATN-1, LCT C/T-13910, and VDR BsmI gene mutations were analyzed with real-time PCR. We did not detect a statistically significant difference between AIS and control groups in respect to those three different gene polymorphisms (p < 0.05). We next evaluated the associations of all three SNPs with scoliosis curve severity. There was no significant difference between curve severity and gene polymorphisms (p < 0.05). In terms of gene polymorphisms, AIS patients with a family history of AIS did not significantly differ from AIS patients who did not have history (p < 0.05). AIS might be caused by many different gene mutations, biomechanical mechanisms that have been modified by environmental factors, different biological interactions, modulation of growth, or a synergy of different factors causing abnormal control of growth. However, the existing knowledge is still not enough to explain the etiopathogenesis of AIS.
Read more
Characterization of arthroplasty tissue after 14 years post-cup arthroplasty: A morphological and biochemical assessment
Richard D Coutts
David Amiel
Frederick L. Harwood
Wayne H. Akeson
The cup arthroplasty has been reported to cause the formation of a fibrocartilaginous joint surface, which may result in a painless, functional joint. The joint surface of a 38-year-old man with a failed cup arthroplasty implanted for 14 years was examined histologically and biochemically. The joint surface tissue of this patient resembled fibrous connective tissue, with major types of collagen being Type I and Type III. No evidence of cartilaginous transformation in the healing scar was demonstrated, despite several years of successful functioning of the cup arthroplasty.
Read more
Proteomic Identification of 14-3-3ϵ as a Linker Protein Between pERK1/2 Inhibition and BIM Upregulation in Human Osteosarcoma Cells.
Kyung Ok Kim
Anny Hsu
Heon Goo Lee
Francis Young-In Lee
Despite advancements in multimodality chemotherapy, conventional cytotoxic treatments still remain ineffective for a subset of patients with aggressive metastatic or multifocal osteosarcoma. It has been shown that pERK1/2 inhibition enhances chemosensitivity to doxorubicin and promotes osteosarcoma cell death in vivo and in vitro. One of the pro-apoptotic mechanisms is upregulation of Bim by pERK1/2 inhibitors. To this end, we examined proteomic changes of 143B human osteosarcoma cells with and without treatment of PD98059, pERK1/2 inhibitor. Specifically, we identified 14-3-3ϵ protein as a potential mediator of Bim expression in response to inhibition of pERK1/2. We hypothesized that 14-3-3ϵ mediates upregulation of Bim expression after pERK1/2 inhibition. We examined the expression of Bim after silencing 14-3-3ϵ using siRNA. The 14-3-3ϵ gene silencing resulted in downregulation of Bim expression after PD98059 treatment. These data indicate that 14-3-3ϵ is required for Bim expression and that it has an anti-cancer effect under pERK1/2 inhibition in 143B cells. By playing an essential role upstream of Bim, 14-3-3ϵ may potentially be a coadjuvant factor synergizing the effect of pERK1/2 inhibitors in addition to conventional cytotoxic agents for more effective osteosarcoma treatments. © 2014 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res.
Read more
Pentadecapeptide BPC 157 (PL 14736) Improves Ligament Healing in the Rat
Tomislav Cerovecki
Ivan Bojanic
Luka Brcic
Predrag Sikiric
We improved medial collateral ligament (MCL) healing throughout 90 days after surgical transection. We introduced intraperitoneal, per-oral (in drinking water) and topical (thin cream layer) peptide therapy always given alone, without a carrier. Previously, as an effective peptide therapy, stable gastric pentadecapeptide BPC 157 (GEPPPGKPADDAGLV, an anti-ulcer peptide effective in inflammatory bowel disease therapy (PL 14736)) particularly improved healing of transected tendon and muscle and wound healing effect including the expression of the early growth response 1 (egr-1) gene. After MCL transection BPC 157 was effective in rats when given once daily intraperitoneally (10 microg or 10 ng/kg) or locally as a thin layer (1.0 microg dissolved in distilled water/g commercial neutral cream) at the site of injury, first application 30 min after surgery and the final application 24 h before sacrifice. Likewise, BPC 157 was effective given per-orally (0.16 microg/ml in the drinking water (12 ml/day/rat)) until sacrifice. Commonly, BPC 157 microg-ng-rats exhibited consistent functional, biomechanical, macroscopic and histological healing improvements. Thus, we suggest BPC 157 improved healing of acute ligament injuries in further ligament therapy.
Read more
The (−1486T/C) promoter polymorphism of the TLR-9 gene is associated with end-stage knee osteoarthritis in a Chinese population
Sui-Lung Su
Hsin-Yi Yang
Chian-Her Lee
Herng-Sheng Lee
Based on the recent observation that Toll-like receptors (TLRs) may be involved in the pathogenesis of osteoarthritis (OA) we explored the possibility that human TLR gene polymorphisms are associated with OA. Two separate populations were studied in a two-stage case-control study with a total of 503 OA patients and 428 healthy controls. The TLR-2, TLR-4, and TLR-9 genotypes were assessed by real-time polymerase chain reaction. Our data demonstrated a lack of association among TLR-2, TLR-4, and TLR-9 (T-1237C) polymorphisms and the risk of developing OA in both stages of the study. T-1486C was significantly associated with OA in both populations with G1635A of TLR-9 gene was found to be significantly associated with OA when the two populations were combined. Stratifying the samples by K-L score there were significant differences in the genotype of the TLR-9 T-1486C and G1635A between OA of the knee grade 4 and controls. In haplotype analyses, the haplotype TTG and TTA revealed higher risk of OA and TCA confers a lower risk of OA in combined population. The present results demonstrate that TLR-9 polymorphisms, in particular T-1486C is significantly associated with OA. TLR-9 gene polymorphisms may play a role in the etiology of knee OA.
Read more
Hannafin JA, Attia EA, Henshaw R, Warren RF, Bhargava MM.. Effect of cyclic strain and plating matrix on cell proliferation and integrin expression by ligament fibroblasts. J Orthop Res 24: 149-158
Jo A Hannafin
Erik Attia
Ross Henshaw
Madhu M Bhargava
The role of cell surface integrins in cell migration, proliferation, and attachment to matrix molecules is well known. Integrin-matrix interactions have been implicated in mechanotransduction and load transmission from the outside to the inside of the cell. In this study, the effect of cyclic strain on the cell proliferation, attachment, and expression of integrin subunits beta1, beta3, and alpha5 was determined in anterior cruciate ligament (ACL) and medial collateral ligament (MCL) fibroblasts grown on polystyrene, Type I collagen, laminin, elastin, and fibronectin. ACL fibroblast proliferation was not affected by growth substrate whereas MCL cells reached confluence more rapidly on fibronectin compared with collagen or polystyrene. Exposure to 5% cyclic strain resulted in a significant decrease in ACL and MCL fibroblast proliferation on fibronectin and Type I collagen. MCL cells showed a greater strain-dependent inhibition of cells grown on a fibronectin substrate than those grown on collagen. This matrix-dependent effect of strain on cell proliferation was not seen with ACL cells. Attachment of ACL and MCL fibroblasts was stronger to fibronectin compared with Type I collagen, laminin, and polystyrene. In the absence of applied load, the expression of beta1, beta3, and alpha5 subunits was not substrate dependent and the expression of beta1 and alpha5 integrin subunits was higher in MCL cells than ACL cells on all substrates. In contrast, the expression of beta3 integrin subunit was higher in ACL cells than MCL cells. In response to 5% strain, beta1, and alpha5 expression increased in all fibroblasts with MCL cells having a higher magnitude of expression. beta3 expression showed a 90% increase in response to load when grown on laminin for both MCL and ACL fibroblasts and demonstrated no change in expression on Type I collagen or fibronectin. The duration of applied strain from 2 versus 22 h had no effect on cell proliferation or integrin expression.
Read more
Wear and Migration of Highly Cross-Linked and Conventional Cemented Polyethylene Cups with Cobalt Chrome or Oxinium Femoral Heads: A Randomized Radiostereometric Study of 150 Patients
Thomas Kadar
Geir Hallan
Arild Aamodt
Ove Furnes
This randomized study was performed to compare wear and migration of five different cemented total hip joint articulations in 150 patients. The patients received either a Charnley femoral stem with a 22.2 mm head or a Spectron EF femoral stem with a 28 mm head. The Charnley articulated with a γ-sterilized Charnley Ogee acetabular cup. The Spectron EF was used with either EtO-sterilized non-cross-linked polyethylene (Reflection All-Poly) or highly cross-linked (Reflection All-Poly XLPE) cups, combined with either cobalt chrome (CoCr) or Oxinium femoral heads. The patients were followed with repeated RSA measurements for 2 years. After 2 years, the EtO-sterilized non-cross-linked Reflection All-Poly cups had more than four times higher proximal penetration than its highly cross-linked counterpart. Use of Oxinium femoral heads did not affect penetration at 2 years compared to heads made of CoCr. Further follow-up is needed to evaluate the benefits, if any, of Oxinium femoral heads in the clinical setting. The Charnley Ogee was not outperformed by the more recently introduced implants in our study. We conclude that this prostheses still represents a standard against which new implants can be measured.
Read more
Effective therapy of transected quadriceps muscle in rat: Gastric pentadecapeptide BPC 157
Staresinic Mario
Igor Petrovic
Tomislav Novinscak
Predrag Sikiric
We report complete transection of major muscle and the systemic peptide treatment that induces healing of quadriceps muscle promptly and then maintains the healing with functional restoration. Initially, stable gastric pentadecapeptide BPC 157 (GEPPPGKPADDAGLV, M.W. 1419, PL-10, PLD-116, PL 14736 Pliva, Croatia; in trials for inflammatory bowel disease; wound treatment; no toxicity reported; effective alone without carrier) also superiorly accelerates the healing of transected Achilles tendon. Regularly, quadriceps muscle completely transected transversely 1.0 cm proximal to patella presents a definitive defect that cannot be compensated in rat. BPC 157 (10 microg, 10 ng, 10 pg/kg) is given intraperitoneally, once daily; the first application 30 min posttransection, the final 24 h before sacrifice. It consistently improves muscle healing throughout the whole 72-day period. Improved are: (i) biomechanic (load of failure increased); (ii) function (walking recovery and extensor postural thrust/motor function index returned toward normal healthy values); (iii) microscopy/immunochemistry [i.e., mostly muscle fibers connect muscle segments; absent gap; significant desmin positivity for ongoing regeneration of muscle; larger myofibril diameters on both sides, distal and proximal (normal healthy rat-values reached)]; (iv) macroscopic presentation (stumps connected; subsequently, atrophy markedly attenuated; finally, presentation close to normal noninjured muscle, no postsurgery leg contracture). Thus, posttransection healing-consistently improved-may suggest this peptide therapeutic application in muscle disorders.
Read more
Gastric pentadecapeptide BPC 157 accelerates healing of transected rat Achilles tendon and in vitro stimulates tendocytes growth
Staresinic Mario
B Sebecic
Leonardo Patrlj
Predrag Sikiric
In studies intended to improve healing of transected Achilles tendon, effective was a stable gastric pentadecapeptide BPC 157 (GEPPPGKPADDAGLV, M.W. 1419). Currently in clinical trials for inflammatory bowel disease (PLD-116, PL 14736, Pliva), it ameliorates internal and external wound healing. In rats, the right Achilles tendon transected (5 mm proximal to its calcaneal insertion) presents with a large tendon defect between cut ends. Agents (/kg b.w., i.p., once time daily) (BPC 157 (dissolved in saline, with no carrier addition) (10 microg, 10 ng or 10 pg) or saline (5.0 ml)), were firstly applied at 30 min after surgery, the last application at 24 h before autopsy. Achilles functional index (AFI) was assessed once time daily. Biomechanical, microscopical and macroscopical assessment was on day 1, 4, 7, 10 and 14. Controls generally have severely compromised healing. In comparison, pentadecapeptide BPC 157 fully improves recovery: (i) biomechanically, increased load of failure, load of failure per area and Young's modulus of elasticity; (ii) functionally, significantly higher AFI-values; (iii) microscopically, more mononuclears and less granulocytes, superior formation of fibroblasts, reticulin and collagen; (iv) macroscopically, smaller size and depth of tendon defect, and subsequently the reestablishment of full tendon integrity. Likewise, unlike TGF-beta, pentadecapeptide BPC 157, presenting with no effect on the growth of cultured cell of its own, consistently opposed 4-hydroxynonenal (HNE), a negative modulator of the growth. HNE-effect is opposed in both combinations: BPC 157+HNE (HNE growth inhibiting effect reversed into growth stimulation of cultured tendocytes) and HNE+BPC 157(abolished inhibiting activity of the aldehyde), both in the presence of serum and serum deprived conditions. In conclusion, these findings, particularly, Achilles tendon transection fully recovered in rats, peptide stability suitable delivery, usefully favor gastric pentadecapeptide BPC 157 in future Achilles tendon therapy.
Read more
Achilles Detachment in Rat and Stable Gastric Pentadecapeptide BPC 157: Promoted Tendon-to-Bone Healing and Opposed Corticosteroid Aggravation
Andrija Krivic
Tomislav Anic
Sven Seiwerth
Predrag Sikiric
Stable gastric pentadecapeptide BPC 157 (BPC 157, as an antiulcer agent in clinical trials for inflammatory bowel disease; PLD-116, PL 14736, Pliva, no toxicity reported) alone (without carrier) ameliorates healing of tendon and bone, respectively, as well as other tissues. Thereby, we focus on Achilles tendon-to-bone healing: tendon to bone could not be healed spontaneously, but it was recovered by this peptide. After the rat's Achilles tendon was sharply transected from calcaneal bone, agents [BPC 157 (10 microg, 10 ng, 10 pg), 6alpha-methylprednisolone (1 mg), 0.9% NaCl (5 mL)] were given alone or in combination [/kg body weight (b.w.) intraperitoneally, once time daily, first 30-min after surgery, last 24 h before analysis]. Tested at days 1, 4, 7, 10, 14, and 21 after Achilles detachment, BPC 157 improves healing functionally [Achilles functional index (AFI) values substantially increased], biomechanically (load to failure, stiffness, and Young elasticity modulus significantly increased), macro/microscopically, immunohistochemistry (better organization of collagen fibers, and advanced vascular appearance, more collagen type I). 6alpha-Methylprednisolone consistently aggravates the healing, while BPC 157 substantially reduces 6alpha-methylprednisolone healing aggravation. Thus, direct tendon-to-bone healing using stabile nontoxic peptide BPC 157 without a carrier might successfully exchange the present reconstructive surgical methods.
Read more
Letter to the editor Re: Wei, Li-Cheng et al. J Orthop Res 30:1572-1576 2012
Guanghua Lei
Letter to the editor Re: Wei, Li-Cheng e;t al. J Orthop Res 30:1572-1576 2012
James D Heckman
Kevin F. Spratt
Proprioception after an acute knee ligament injury: A longitudinal study on 16 consecutive patients
Thomas Fridén
David Roberts
Rose Zätterström
Ulrich Moritz
A defect in proprioception has been found in selected patient groups that have an anterior cruciate ligament deficient knee at different times after the original injury. The time of development and the extent of such defects were studied longitudinally on 16 consecutive patients. During the first year after a primary knee injury, which included a complete rupture of the anterior cruciate ligament, we repeatedly performed three tests of proprioception: (a) one to determine the threshold for detecting a passive motion from starting positions of 20 and 40 degrees, (b) an active reproduction of a passive angular change, and (c) a visual estimation of a passive angular change. The injured limb was compared with the uninjured limb and with the limbs of an age-matched reference group of healthy subjects. The population did not have a normal distribution, and some patients had consistently extreme recordings in the threshold tests at the various times of testing. Significant differences were found between the groups at the starting position of 20 degrees, when the injured knee was compared with the uninjured knee, after 1 month (p = 0.05), and after 2 months (p = 0.03). There was a trend toward a higher threshold for detecting a passive motion when the injured side was compared with the knees of the reference group at 1 month (p = 0.06) but not later on. A similar pattern was found for the injured knee at the starting position of 40 degrees, but it was not significant. An impaired ability to detect a passive motion was registered for the nearly extended knee 1 and 2 months after a primary injury. In the active reproduction and visual estimation tests, no significant defects were found at any time during the first year in these consecutively studied patients.
Read more
Spinal mobility and posture in 8- to 16-year-old children
Guy Mellin
Mikko Poussa
Spinal mobility and posture were measured in 294 8-16-year-old boys and girls, divided into five age groups. The upper thoracic sagittal alignment was more vertical among girls, but the postural curves showed no significant age-related differences for either sex. Among both boys and girls thoracic extension, lateral flexion, and rotation decreased significantly between the ages of 12 and 13, but with the exception of extension they returned to the previous level by age 16. Girls were significantly different from boys at 13 years of age. In the thoracic spine, girls had less kyphosis, and were stiffer in forward and lateral flexion, with more rotation to the right than to the left. In the lumbar spine, lateral flexion increased after the age of 10 in both sexes. Between the ages of 8 and 14 lumbar lateral flexion was significantly greater among girls than among boys, whereas extension and rotation was greater only at the ages of 8 and 10 years. With increasing age, a shift from left to right dominance in lumbar lateral flexion was found in girls only.
Read more
Inducible displacements of cemented tibial components during weight-bearing and knee extension. Observations during dynamic radiostereometry related to joint positions and 2 years history of migration in 16 TKR
Johan Uvehammer
Johan Kärrholm
The inducible displacements of the tibial component caused by active extension were studied in 16 knees 1 yr after an AMK total knee arthroplasty with either flat. concave or posterior-stabilised (PS) designs of the joint area. Continuous change of the position of the tibial component occurred with proceeding extension. Rocking, subsidence and lift-off at different localisations were observed. In 3 of 4 knees with flat inserts the tibial component tilted anteriorly from 45 degrees to 35 degrees of flexion. A similar anterior tilt was seen in 2 of 6 with concave inserts and 5 of 6 with the PS design, but the tilting started later, when the knee had 5-20 degrees more extension. From 45-15 degrees of flexion most components tilted into valgus. Three knees (1 concave with, 1 concave without PCL and 1 PS) showed a sudden tilt into varus direction followed by a rocking motion in the opposite direction. The other types of displacements studied showed a more uniform pattern. The inducible maximum translation (MTPM) at 20 degrees of extension tended to be associated with increased migration between 0 and 2 yr when measured with the same parameter (Spearman's rho = 0.54, P = 0.03). Increased medial displacement of the center of the proximal tibia at 25 degrees was associated with increased anterior tilt. This type of motion was most commonly seen with the concave design. Our observations demonstrate that the forces acting on the tibial component vary during active extension, which results in rocking movements. This will influence the migration and the patttern of wear, factors of importance for the clinical longevity of a total knee replacement (TKR).
Read more
Treatment of antigen-induced arthritis in rabbits with dysprosium-165-ferric hydroxide macroaggregates
Joseph D. Zuckerman
Clement B. Sledge
Sonya Shortkroff
Prasanna Venkatesan
Dysprosium-165-ferric hydroxide macroaggregates (165Dy-FHMA) was used as an agent of radiation synovectomy in an antigen-induced arthritis model in New Zealand white rabbits. Animals were killed up to 6 months after treatment. 165Dy-FHMA was found to have a potent but temporary antiinflammatory effect on synovium for up to 3 months after treatment. Treated knees also showed significant preservation of articular cartilage architecture and proteoglycan content compared with untreated controls, but only during the first 3 months after treatment. In animals killed 3 and 6 months after treatment there were only minimal differences between the treated and untreated knees, indicating that the antiinflammatory effects on synovial tissue and articular cartilage preservation were not sustained.
Read more
rhBMP-2, rhVEGF(165), rhPTN and thrombin-related peptide, TP508 induce chemotaxis of human osteoblasts and microvascular endothelial cells
Xian Liu
Yuxin Cui
Lisa McIlmurray
Hali Wang
Osteogenesis and angiogenesis are inter-linked and tightly regulated processes involved in growth, repair, and bone remodeling. Bone morphogenic protein 2 (BMP-2), vascular endothelial growth factor (VEGF), pleiotrophin (PTN) and thrombin-related peptide, TP508 have all been found to have the ability to promote bone fracture healing by enhancing both the osteogenesis and angiogenesis processes. One of the underlying mechanisms proposed is that mediators for osteogenesis may also be involved in mediating angiogenesis and vice versa. The aim of this study was to examine the chemotactic effects of rhBMP-2, rhVEGF(165), rhPTN and TP508 on human osteoblasts and endothelial cells. Using a direct-viewing chemotaxis assay system, we report for the first time, the direct quantitative observation of chemotaxis of both human osteoblastc cells and microvascular endothelial cells towards sources of rhBMP-2, rhVEGF(165), rhPTN and TP508. This study confirmed that rhBMP-2, rhVEGF(165), rhPTN and TP508 have chemotactic effects on both human osteoblastic and endothelial cells, indicating that these factors are directly involved in promoting angiogenesis and osteogenesis by recruiting osteoblasts and endothelial cells via chemotaxis.
Read more
Goater, JJ, O'Keefe, RJ, Rosier, RN, Puzas, JE and Schwarz, EM. Efficacy of ex vivo OPG gene therapy in preventing wear debris induced osteolysis. J Orthop Res 20: 169-173
J Jeffrey Goater
Regis J O'Keefe
Randy N Rosier
Edward Schwarz
Aseptic loosening of prosthetic implants remains a serious orthopaedic problem and the greatest limitation to total joint arthroplasty. Central to the etiology of aseptic loosening is periprosthetic osteolysis at the bone-implant interface, which is caused by wear debris-induced inflammation. This inflammation produces the critical osteoclast differentiation factor RANKL, which directly stimulates osteoclastogenesis and osteoclastic bone resorption. A dominant factor known to counteract this process is the natural RANKL receptor antagonist protein OPG. Here we explore the potential of ex vivo OPG gene therapy for aseptic loosening by evaluating the eflicacy of stably transfected fibroblast-like synoviocytes (FLS) expressing OPG in preventing wear debris-induced osteoclastogenesis, in a mouse calvaria model. Although the stably transfected fibroblasts produced small amounts of OPG (0.3 ng/ml/72 h/10(6) cells), this protein was very effective in preventing osteoclastic resorption as determined in a bone wafer assay. More importantly. implantation of 10(7) FLS-OPG, together with 30 mg of Ti wear debris, onto the calvaria of mice, completely inhibited osteoclastogenesis 3 days after surgery. Animals given FLS-LacZ control cells, which persisted for 3 days as determined by X-gal staining, together with the Ti particles, had a 6-fold increase in osteoclastogenesis compared to controls without Ti. This increased osteoclastogenesis was completely inhibited by the FLS-OPG, as osteoclast numbers in the calvaria of these animals were similar to that seen in the SHAM controls.
Read more
Absence of linkage to chromosomes 6q and 16p in a Greek population with knee Osteoarthritis
Aspasia Tsezou
Theofilos Karachalios
Pelagia Fytili
Konstantinos Malizos
Osteoarthritis (OA) is a common age-related debilitating disease of the joints characterized by degeneration of the articular cartilage which leads to joint pain, discomfort, and immobility. Several risk factors have been associated with OA including a genetic predisposition. Specific chromosomal regions have thus far been associated with susceptibility to OA, the strongest being on chromosomes 2, 6, and 16. We hereby report our data on 34 Central Greek knee OA families that were investigated for linkage to the chromosome 6q and 16p susceptibility regions. All affected members had undergone total knee replacement surgery (TKR) at a single large Orthopedics Unit in Central Greece. Nineteen microsatellite markers were selected, 15 for chromosome 6q and 4 for chromosome 16p at a distance of approximately 7 cm. Allele fragment sizes were determined by an automated DNA sequencer using the Fragment Analysis Software. Our results revealed a statistically significant difference in the ratio of affected females to males with knee OA and also showed that there is no evidence of linkage to regions 6q and 16p in a cohort of Central Greek pedigrees with knee OA.
Read more
Pulsed electromagnetic fields affect phenotype and connexin 43 protein expression in MLO-Y4 osteocyte-like cells and ROS 17/2.8 osteoblast-like cells
Christoph H Lohmann
Zvi Schwartz
Y Liu
Barbara D Boyan
Osteocytes, the predominant cells in bone, are postulated to be responsible for sensing mechanical and electrical stimuli, transducing signals via gap junctions. Osteocytes respond to induced shear by increasing connexin 43 (Cx43) levels, suggesting that they might be sensitive to physical stimuli like low-frequency electromagnetic fields (EMF). Immature osteoblasts exhibit decreased intercellular communication in response to EMF but no change in Cx43. Here, we examined long term effects of pulsed EMF (PEMF) on MLO-Y4 osteocyte-like cells and ROS 17/2.8 osteoblast-like cells. In MLO-Y4 cell cultures, PEMF for 8 h/day for one, two or four days increased alkaline phosphatase activity but had no effect on cell number or osteocalcin. Transforming growth factor beta-1 (TGF-beta 1) and prostaglandin E(2) were increased, and NO(2-) was altered. PEMFs effect on TGF-beta1 was via a prostaglandin-dependent mechanism involving Cox-1 but not Cox-2. In ROS 17/2.8 cells, PEMF for 24, 48 or 72 h did not affect cell number, osteocalcin mRNA or osteocalcin protein. PEMF reduced Cx43 protein in both cells. Longer exposures decreased Cx43 mRNA. This indicates that cells in the osteoblast lineage, including well-differentiated osteoblast-like ROS 17/2.8 cells and terminally differentiated osteocyte-like MLO-Y4 cells, respond to PEMF with changes in local factor production and reduced Cx43, suggesting decreased gap junctional signaling.
Read more
Attachment of epiphyseal cartilage cells and 17/28 rat osteosarcoma osteoblasts using mussel adhesive protein
John P Fulkerson
Louis Norton
Gloria Gronowicz
Carl W Nissen
These experiments show that mussel adhesive protein (MAP) enhances the attachment of osteoblasts and epiphyseal cartilage cells to plastic culture dishes and Vitallium. When MAP was applied to culture plate surfaces, there were two- to fivefold increases in the numbers of cells attaching compared to control surfaces (no MAP). Results were confirmed using two different cell attachment assay techniques. Osteoblast replication and culture on MAP is possible, suggesting that MAP is not toxic to cells. MAP also holds applied cells to surfaces as initially attached.
Read more
Interleukin-17 synergizes with IFNIγ or TNFα to promote inflammatory mediator release and intercellular adhesion molecule-1 (ICAM-1) expression in human intervertebral disc cells
Mostafa A Gabr
Liufang Jing
Antonia R Helbling
Jun Chen
Interleukin-17 (IL-17) is a cytokine recently shown to be elevated, along with interferon-γ (IFNγ) and tumor necrosis factor (TNFα), in degenerated and herniated intervertebral disc (IVD) tissues, suggesting a role for these cytokines in intervertebral disc disease. The objective of our study was to investigate the involvement of IL-17 and costimulants IFNγ and TNFα in intervertebral disc pathology. Cells were isolated from anulus fibrosus and nucleus pulposus tissues of patients undergoing surgery for intervertebral disc degeneration or scoliosis. The production of inflammatory mediators, nitric oxide (NOx), prostaglandin E2 (PGE2) and interleukin-6 (IL-6), as well as intercellular adhesion molecule (ICAM-1) expression, were quantified for cultured cells following exposure to IL-17, IFNγ, and TNFα. Intervertebral disc cells exposed to IL-17, IFNγ, or TNFα showed a remarkable increase in inflammatory mediator release and ICAM-1 expression (GLM and ANOVA, p < 0.05). Addition of IFNγ or TNFα to IL-17 demonstrated a synergistic increase in inflammatory mediator release, and a marked increase in ICAM-1 expression. These findings suggest that IVD cells not only respond with a catabolic phenotype to IL-17 and costimulants IFNγ and TNFα, but also express surface ligands with consequent potential to recruit additional lymphocytes and immune cells to the IVD microenvironment. IL-17 may be an important regulator of inflammation in the IVD pathologies.
Read more
Trunk positioning accuracy in children 7-18 years old
James Ashton-Miller
Kelly M. McGlashen
Albert B. Schultz
Trunk proprioception was measured in 253 healthy children 7-18 years of age using infrared markers placed on the back of the head and on the skin over the T1, T8, and S1 spinous processes. The children were tested for their accuracy in sensing return of the head and trunk to a centered, neutral position in the frontal plane. Whole-body sway was also quantified during 10 s of relaxed standing by measuring mean amplitudes of trunk marker and foot center of pressure (CP) movements. The results show that trunk positioning accuracy improved significantly with age (p = 0.000). Subjects could position their trunk in the frontal plane to within a mean (+/- SD) of 2.5 (+/- 1.1) and 0.9 (+/- 0.6) degrees of the neutral position at ages 7 and 18 years, respectively. No statistically significant gender differences were found. At every age trunk positioning accuracy was diminished in the presence of a continuous external trunk moment (equivalent to 0.01 x body weight x height), although not significantly so. Neither mean trunk sway nor CP amplitudes were significantly correlated with age or sex. The overall results suggest that spine decompensation is only abnormal when it exceeds 20 mm in healthy children and adolescents.
Read more
AAOS/NIH/ORS workshop. Injury and repair of the musculoskeletal soft tissues. Savannah, Georgia, June 18-20, 1987
Savio L-Y. Woo
Joseph Addision Buckwalter
Top-cited authors
Benjamin J Fregly
Rice University
Farshid Guilak
Washington University in St. Louis
Thor Besier
University of Auckland
Scott L Delp
Stanford University
James C Iatridis
Icahn School of Medicine at Mount Sinai
About us
Help Center
Business solutions
© 2008-2022 ResearchGate GmbH. All rights reserved.
Discover by subject area
Join for free
Log in


text:
Sizing Guide for Shrimp
|Common Sizing Term (Varies)
|Shrimp Count Per Pound
|Approx. Count Per 3 oz Serving
|Extra Large
|6-7 shrimp
|8-9 shrimp
|Medium Large
|9-10 shrimp
|10-12 shrimp
8 •
How many shrimp are in a pound of shrimp?
The result is that every pound of shrimp includes between 8 and 12 large, fresh shrimp. In fact, giant shrimp are the biggest shrimp available on the market! Our somewhat smaller wild caught shrimp are in the jumbo range, with 16 to 20 shrimp per pound (U-16/20) and are available in a variety of sizes.
How many jumbo shrimp are in a half pound?
Meanwhile, jumbo shrimp are branded as ' 21/25,' which implies you'll get between 21 and 25 shrimp per pound when you buy them.
How many Oz is 15 medium shrimp?
Food Lion – Medium-Range Cooked Shrimp ( 3 Oz., 15 Pieces)
Is 1 pound of shrimp enough?
Purchasing Suggestions for Shrimp When purchasing shrimp, the general rule of thumb is that you should purchase 1 pound of raw, unpeeled shrimp per person, or 1/2 -1/3 pound of cooked, peeled shrimp per person if you are purchasing it cooked and peeled. The amount of shrimp included in a pound will vary depending on the size of the shrimp.
How many shrimp come in a half pound?
In the United States, shrimp are sold by the pound, rather than by the count. The number on the label will show the number of shrimp that are included within the container. If the number is
21/25, you know that there should be between 21 and 25 shrimp in each pound of shrimp.
What is the difference in shrimp sizes?
Medium and smaller shrimp (about 9 to 11 shrimp per person). Large to large shrimp (about 5 to 7 shrimp per person). Excessive jumbo and above, around 3 to 5 per person
What is the best size shrimp for a shrimp boil?
Choose Large Shrimp, which average out to about 30 per pound on average, if you're making boiling shrimp. They present well on a dish and are substantial enough to serve as a main course on its own. If you're boiling the shrimp for a meal, buy at least two pounds; if you're hosting a party, buy at least four pounds.
Can you eat too much shrimp?
Shrimps are well-known for raising LDL cholesterol levels. Specifically, it is this cholesterol that has the potential to cause plaque to build up in your arteries, which can ultimately lead to a heart attack or stroke. As a result, if you consume an excessive amount of shrimp, you are significantly raising your LDL cholesterol levels and putting yourself at unneeded
What is the average size of a shrimp?
Shrimp varies in size from a few millimetres to more than 20 centimetres (about 8 inches); the typical size is between 4 and 8 centimetres (1.5 to 3 inches). Prawns are commonly used to refer to those who are larger in size.
How do you measure Shrimp in ounces?
From a few millimetres to more than 20 cm (about 8 inches) in length, shrimp are available in a variety of sizes. The typical size is between 4 and 8 cm (1.5 to 3 inches). Prawns are a term used to describe larger persons.
How many calories are in 15 medium Shrimp?
In a serving of 15 medium shrimp, there are 95 calories.
How do you measure shrimp?
Shrimp are sized in the United States according to their weight, and more precisely, how many shrimp are included in one pound: Increasingly large sizes are denoted by the letter U in front of the integer (for example, U/10). The letter U stands for 'under,' which means that one pound of shrimp contains less than ten shrimp.
How much shrimp is a serving?
The following are some general guidelines for shrimp serving sizes. A main meal of 6–8 ounces of raw headless shell on shrimp is recommended. 2–3 ounces of uncooked headless shell on shrimp served as an appetizer When serving shrimp for a large gathering, I serve 4 ounces per person if there
are many other dishes on the table to complement the shrimp.

subset: commoncrawl


text:
Login/Register to interact with Peter
Peter Taylor
Professional Speaker - Trainer - Consultant - Faculty lead at #ThePMTribe, Pembury Partners Ltd
Sandhurst, United Kingdom
PMI Chapter:
PMI United Kingdom Chapter
Contact me about: Current employment opportunities,Consulting/freelance opportunities,Career opportunities,Expertise requests,Speaking opportunitiesPeter Taylor is the author of two best-selling books on 'Productive Laziness' – 'The Lazy Winner' and 'The Lazy Project Manager' and is a professional speaker.
More information can be found at www.thelazyprojectmanager.com and #ThePMTribe at www.thepmtribe - come check it out
"Brevity is the soul of lingerie."
- Dorothy Parker


text:
The District Court of Frankfurt am Main has – in a decision that was criticized early on – stated that in the case of red light violations with a so-called Sport Utility Vehicle (SUV), an increase in the standard fine may be appropriate (District Court of Frankfurt am Main, judgment of June 3rd, 2022, 974 OWi 533 Js-OWi 18474/22).
According to the findings of the court in fine proceedings, the person concerned drove his vehicle in Frankfurt am Main (crossing Friedrich-Ebert-Anlage), a so-called BMW Sport Utility Vehicle (SUV for short), which in terms of its design differed from normal motor vehicles in such that it has increased ground clearance, enter the crossing area regulated by the traffic light system. At this point, the red phase had already lasted longer than 1.1 seconds.
The district court, which also determined the correctness of the measurement made using a permanently installed measuring column, saw an increase in the standard fine provided for this by the applicable catalog of fines as necessary due to the special vehicle procurement in the specific case.
This is justified by the increased operating risk of the motor vehicle used, whose boxy design and raised front end increase the risk of injury to other road users:
In the assessment, the court took into account the fact that the person concerned had several previous entries in the register of fitness to drive within the framework of Section 3 (1) BKatV. Accordingly, the fine had to be increased.
In addition, the increased operating risk of the vehicle used was taken into account when assessing the fine at the expense of the person concerned. The boxy design and increased front end of the vehicle due to the greater ground clearance increase the risk of injury to other road users in an SUV. There is therefore an increased operating risk compared to a passenger car of conventional design (cf. also Higher Regional Court Hamm, judgment of September 30, 1996 – 6 U 63/96, NZV 1997, 230).
Due to the greater abstract danger from the motor vehicle being driven, the red light violation committed is more serious than the normal case; especially since the provisions of § 37 StVO on alternating light signals aim to protect crossing road users in the intersection area of the traffic light system in the event of a collision. Therefore, this case has a special feature that distinguishes it from ordinary circumstances, so that the standard fine must be increased accordingly.
Against the background of these considerations, the court considers that a Fine of 350 euros to fix.
Due to the greater abstract danger posed by the vehicle involved in the crime, according to the District Court of Frankfurt am Main, the red light violation committed is more serious than the normal case. This applies in particular to the objective of §37 StVO on alternating light signals, which protects crossing road users in the area of traffic lights intended in a collision (source: press release of the court)


text:
|Chet Ramey
|Re: [Help-bash] set other pager for TAB completion possibilities
|Sat, 15 Mar 2014 14:16:24 -0400
|Mozilla/5.0 (Macintosh; Intel Mac OS X 10.9; rv:24.0) Gecko/20100101 Thunderbird/24.3.0
On 3/15/14, 5:45 AM, Christof Warlich wrote: > Hi, > > though I fear that it is just not possible, I'd like to confirm: > > Wen I press TAB twice to see the autocompletion possibilities of a command, > and the list of choices is big, bash asks if it should display all > possibilities. Saying yes, bash seems to use the pager "more" to display > the possibilities. Readline does not use `more', or any other external program; it uses its own internal pager. There is no user-configurable hook to change that. Chet -- ``The lyf so short, the craft so long to lerne.'' - Chaucer ``Ars longa, vita brevis'' - Hippocrates Chet Ramey, ITS, CWRU address@hidden http://cnswww.cns.cwru.edu/~chet/
|[Prev in Thread]
|Current Thread
|[Next in Thread]


text:
June 24, 2014
Legislative Assembly - Province of British Columbia
VICTORIA – The Special Committee to Review the
Personal Information Protection Act today launched a public consultation on the
Personal Information Protection Act (RSBC 2003, c. 63). The act governs how private sector organizations can collect, use and disclose personal information. It also requires organizations to protect and secure personal information against unauthorized use or disclosure, and grants individuals the right to access their own personal information.
"The committee is undertaking a comprehensive review of B.C.'s private sector privacy legislation," said Committee Chair Mike Bernier. "We are holding a public consultation to gather important information on how well the act is working, and whether changes are necessary."
"The last review of the act took place in 2008," said Deputy Chair George Heyman. "The committee wants to hear from stakeholders and interested citizens on ways in which the legislation might be updated."
The consultation will include a call for written submissions and public hearings scheduled for Sept. 8 and 9, 2014. To participate, or to learn more about the Act, please visit the Committee website at:
The call for public input closes on Friday, Sept. 19, 2014.
The Special Committee to Review the
Personal Information Protection Act is an all-party committee of the Legislative Assembly of British Columbia. The committee members are:
Mike Bernier, MLA Peace River South (chair);
George Heyman, MLA Vancouver-Fairview (deputy chair);
Donna Barnett, MLA Cariboo-Chilcotin;
Dr. Doug Bing, MLA Maple Ridge-Pitt Meadows;
Simon Gibson, MLA Abbotsford-Mission;
Sue Hammell, MLA Surrey-Green Timbers;
Marvin Hunt, MLA Surrey-Panorama;
Doug Routley, MLA Nanaimo-North Cowichan;
For further Information:
Susan Sourial
Committee Clerk
Room 224, Parliament Buildings
Victoria, B.C. V8V 1X4
Telephone: 250 356-2933
Toll-free: 1-877-428-8337
Fax: 250 356-8172E-mail: