Dataset Information

Targeted designed variants of alpha-2-macroglobulin (A2M) attenuate cartilage degeneration in a rat model of osteoarthritis induced by anterior cruciate ligament transection.

ABSTRACT: The study was performed to evaluate whether targeted alpha-2-macroglobulin (A2M) variants have a similar or enhanced function at wild-type (wt)-A2M to attenuate cartilage degeneration in vivo.In and ex-vivo experiment, bovine cartilage explants (BCE) were incubated with TNF-? and IL-1? with or without wt-A2M or A2M variants. Cartilage catabolism was measured in culture supernatant by sulfated glycosaminoglycan (sGAG). In an in-vivo experiment, 2-month-old male Wistar rats (n?=?77) were randomly divided into seven groups and treated with different doses of A2M or its variants by intra-articular injection at 24 hours and day 14 after anterior cruciate ligament transection (ACLT), receiving (1) ACLT/PBS; (2) ACLT/wt-A2M (0.153 mg); (3) ACLT/CYT-108 A2M (0.153 mg); (4) ACLT/CYT-108 A2M (0.077 mg); (5) ACLT/CYT-98 A2M (0.153 mg); (6) ACLT/CYT-98 A2M (0.077 mg); or (7) sham/PBS. The joints and synovial lavage were collected 8 weeks after surgery. Fluorescence molecular tomography was used to monitor inflammation in vivo using probes ProSense and MMPSense at 24 hours, and weeks 2, 4, and 6 after surgery. The cartilage damage was quantified using Osteoarthritis Research Society International score and matrix metalloproteinase (MMP)-3, -13, collagen (Col) X, Col 2, Runx2, and aggrecan (Acan) were detected by immunohistochemical analysis (IHC), ELISA, and RT-PCR.A2M variants inhibited catabolism in the BCE model by up to 200% compared with wt-A2M. ProSense and MMPSense were dramatically increased in all groups after surgery. Supplemental A2M or its variants reduced ProSense and MMPSense compared with the PBS treatment. Less cartilage damage, lower MMP-13 and Col 2 degraded product, and stronger Col 2 synthesis were detected in animals treated with A2M or its variants compared with PBS-treated animals. A2M and its variants enhanced Col 2 and Acan synthesis, and suppressed MMP-3, MMP-13, Runx2, and Col X production. A2M-108 variant demonstrated less cartilage damage compared with wt-A2M and A2M-98 variant.The targeted variants of A2M have a chondroprotective effect similar to wt-A2M. However, A2M-108 variant has enhanced function to attenuate cartilage degeneration compared with wt-A2M.

SUBMITTER: Zhang Y

PROVIDER: S-EPMC5526282 | biostudies-literature | 2017 Jul

REPOSITORIES: biostudies-literature

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Targeted designed variants of alpha-2-macroglobulin (A2M) attenuate cartilage degeneration in a rat model of osteoarthritis induced by anterior cruciate ligament transection.

Zhang Yang Y Wei Xiaochun X Browning Shawn S Scuderi Gaetano G Hanna Lewis S LS Wei Lei L

Arthritis research & therapy 20170725 1

<h4>Background</h4>The study was performed to evaluate whether targeted alpha-2-macroglobulin (A2M) variants have a similar or enhanced function at wild-type (wt)-A2M to attenuate cartilage degeneration in vivo.<h4>Methods</h4>In and ex-vivo experiment, bovine cartilage explants (BCE) were incubated with TNF-α and IL-1β with or without wt-A2M or A2M variants. Cartilage catabolism was measured in culture supernatant by sulfated glycosaminoglycan (sGAG). In an in-vivo experiment, 2-month-old male ...[more]

PMID: 28743292

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Project description:ObjectiveTo investigate the mitigate efficacy of Chinese medicine Lingzhi (LZ) and San-Miao-San (SMS) combined with hyaluronic acid (HA)-gel in attenuating cartilage degeneration in traumatic osteoarthritis (OA).MethodsThe standardized surgery of anterior cruciate ligament transection (ACLT) was made from the medial compartment of right hind limbs of 8-week-old female SD rats and resulted in a traumatic OA. Rats (n = 5/group) were treated once intra-articular injection of 50 μl HA-gel, 50 μl HA-gel+50 μg LZ-SMS, 50 μl of saline+50 μg LZ-SMS and null (ACLT group) respectively, except sham group. Limbs were harvested for μCT scan and histopathological staining 3-month post-treatment. Inflammatory cytokines from plasma and synovial fluid were detected using Immunology Multiplex Assay kit. The putative targets of active compounds in LZ-SMS and known therapeutic targets for OA were combined to construct protein-protein interaction network. Gene Ontology and Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analysis was adopted to predict the potential targets and signaling pathway of LZ-SMS in OA through the tool of DAVID Bioinformatics.ResultsIn vivo, HA-gel + LZ-SMS treatment resulted in a higher volume ratio of hyaline cartilage (HC)/calcified cartilage (CC) and HC/Sum (total volume of cartilage), compared to ACLT and HA-gel groups. In addition, histological results showed the elevated cartilage matrix, chondrogenic and osteoblastic signals in HA-gel + LZ-SMS treatment. Treatment also significantly altered subchondral bone (SCB) structure including an increase in BV/TV, Tb.Th, BMD, Conn.Dn, Tb.N, and DA, as well as a significant decrease in Tb.Sp and Po(tot), which implied a protective effect on maintaining the stabilization of tibial SCB microstructure. Furthermore, there was also a down-regulated inflammatory cytokines and upregulated anti-inflammatory cytokine IL-10 in HA+LZ-SMS group. Finally, 64 shared targets from 37 active compounds in LZ-SMS related to the core genes for the development of OA. LZ-SMS has a putative role in regulating inflammatory circumstance through influencing the MAPK signaling pathway.ConclusionOur study elucidated a protective effect of HA-gel + LZ-SMS in mitigating cartilage degradation and putative interaction with targets and signaling pathway for the development of traumatic OA.The translational potential of this articleOur results provide a biological rationale for the use of LZ-SMS as a potential candidate for OA treatment.

Project description:BackgroundOsteoarthritis (OA) is a debilitating, progressive joint disease.MethodsSimilar to the disease progression in humans, sequential events of early cartilage degradation, subchondral osteopenia followed by sclerosis, and late osteophyte formation were demonstrated in the anterior cruciate ligament transection (ACLT) or ACLT with partial medial meniscectomy (ACLT + MMx) rat OA models. We describe a reliable and consistent method to examine the time dependent changes in the gene expression profiles in articular cartilage and subchondral bone.ResultsLocal regulation of matrix degradation markers was demonstrated by a significant increase in mRNA levels of aggrecanase-1 and MMP-13 as early as the first week post-surgery, and expression remained elevated throughout the 10 week study. Immunohistochemistry confirmed MMP-13 expression in differentiated chondrocytes and synovial fibroblasts at week-2 and cells within osteophytes at week-10 in the surgically-modified-joints. Concomitant increases in chondrocyte differentiation markers, Col IIA and Sox 9, and vascular invasion markers, VEGF and CD31, peaked around week-2 to -4, and returned to Sham levels at later time points in both models. Indeed, VEGF-positive cells were found in the deep articular chondrocytes adjacent to subchondral bone. Osteoclastic bone resorption markers, cathepsin K and TRAP, were also elevated at week-2. Confirming bone resorption is an early local event in OA progression, cathepsin K positive osteoclasts were found invading the articular cartilage from the subchondral region at week 2. This was followed by late disease events, including subchondral sclerosis and osteophyte formation, as demonstrated by the upregulation of the osteoanabolic markers runx2 and osterix, toward week-4 to 6 post-surgery.ConclusionsIn summary, this study demonstrated the temporal and cohesive gene expression changes in articular cartilage and subchondral bone using known markers of OA progression. The findings here support genome-wide profiling efforts to elucidate the sequential and complex regulation of the disease.

Dataset Information

Targeted designed variants of alpha-2-macroglobulin (A2M) attenuate cartilage degeneration in a rat model of osteoarthritis induced by anterior cruciate ligament transection.

Publications

Targeted designed variants of alpha-2-macroglobulin (A2M) attenuate cartilage degeneration in a rat model of osteoarthritis induced by anterior cruciate ligament transection.

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