Blocking CCN2 Reduces Established Bone Loss Induced by Prolonged Intense Loading by Increasing Osteoblast Activity in Rats
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ABSTRACT:
SUBMITTER: Lambi A
PROVIDER: S-EPMC10494513 | biostudies-literature | 2023 Jun
REPOSITORIES: biostudies-literature
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Project description:The matricellular protein cell communication factor 2/connective tissue growth factor (CCN2/CTGF) is critical to development of neuromuscular fibrosis. Here, we tested whether anti-CCN2 antibody treatment will reduce established forepaw fibro-degenerative changes and improve function in a rat model of overuse injury. Adult female rats performed a high repetition high force (HRHF) task for 18 weeks. Tissues were collected from one subset after 18 wks (HRHF-Untreated). Two subsets were provided 6 wks of rest with concurrent treatment with anti-CCN2 (HRHF-Rest/anti-CCN2) or IgG (HRHF-Rest/IgG). Results were compared to IgG-treated Controls. Forepaw muscle fibrosis, neural fibrosis and entheseal damage were increased in HRHF-Untreated rats, compared to Controls, and changes were ameliorated in HRHF-Rest/anti-CCN2 rats. Anti-CCN2 treatment also reduced phosphorylated-β-catenin (pro-fibrotic protein) in muscles and distal bone/entheses complex, and increased CCN3 (anti-fibrotic) in the same tissues, compared to HRHF-Untreated rats. Grip strength declines and mechanical sensitivity observed in HRHF-Untreated improved with rest; grip strength improved further in HRHF-Rest/anti-CCN2. Grip strength declines correlated with muscle fibrosis, entheseal damage, extraneural fibrosis, and decreased nerve conduction velocity, while enhanced mechanical sensitivity (a pain-related behavior) correlated with extraneural fibrosis. These studies demonstrate that blocking CCN2 signaling reduces established forepaw neuromuscular fibrosis and entheseal damage, which improves forepaw function, following overuse injury.
| S-EPMC10531056 | biostudies-literature
Project description:Tissue fibrosis is a hallmark of overuse musculoskeletal injuries and contributes to functional declines. We tested whether inhibition of CCN2 (cellular communication network factor 2, previously known as connective tissue growth factor, CTGF) using a specific antibody (termed FG-3019 or pamrevlumab) reduces established overuse-induced muscle fibrosis in a clinically relevant rodent model of upper extremity overuse injury. Young adult rats performed a high repetition high force (HRHF) reaching and lever-pulling task for 18 weeks, after first being shaped for 6 weeks to learn this operant task. Rats were then euthanized (HRHF-Untreated), or rested and treated for 6 weeks with FG-3019 (HRHF-Rest/FG-3019) or a human IgG as a vehicle control (HRHF-Rest/IgG). HRHF-Untreated and HRHF-Rest/IgG rats had higher muscle levels of several fibrosis-related proteins (TGFβ1, CCN2, collagen types I and III, and FGF2), and higher muscle numbers of alpha SMA and pERK immunopositive cells, compared to control rats. Each of these fibrogenic changes was restored to control levels by the blocking of CCN2 signaling in HRHF-Rest/FG-3019 rats, as were HRHF task-induced increases in serum CCN2 and pro-collagen I intact N-terminal protein. Levels of cleaved CCN3, an antifibrotic protein, were lowered in HRHF-Untreated and HRHF-Rest/IgG rats, compared to control rats, yet elevated back to control levels in HRHF-Rest/FG-3019 rats. Significant grip strength declines observed in HRHF-Untreated and HRHF-Rest/IgG rats, were restored to control levels in HRHF-Rest/FG-3019 rats. These results are highly encouraging for use of FG-3019 for therapeutic treatment of persistent skeletal muscle fibrosis, such as those induced with chronic overuse.
| S-EPMC7200299 | biostudies-literature
Project description:Purpose/Aim: We recently found that blocking CCN2 signaling using a monoclonal antibody (FG-3019) may be a novel therapeutic strategy for reducing overuse-induced tissue fibrosis. Since CCN2 plays roles in osteoclastogenesis, and persistent performance of a high repetition high force (HRHF) lever pulling task results in a loss in trabecular bone volume in the radius, we examined here whether blocking CCN2 signaling would reduce the early catabolic effects of performing a HRHF task for 3 weeks. Materials and Methods: Young adult, female, Sprague-Dawley rats were operantly shaped to learn to pull at high force levels, before performing the HRHF task for 3 weeks. HRHF task rats were then left untreated (HRHF Untreated), treated in task weeks 2 and 3 with a monoclonal antibody that antagonizes CCN2 (HRHF+FG-3019), or treated with an IgG (HRHF+IgG), while continuing to perform the task. Non-task control rats were left untreated. Results: In metaphyseal trabeculae of the distal radius, HRHF Untreated and HRHF-IgG rats showed increased osteoblast numbers and other indices of bone formation, compared to controls, yet decreased trabecular bone volume, increased osteoclast numbers, and increased serum CTX-1 (a serum biomarker of bone resorption). HRHF+FG-3019 rats also showed increased osteoblast numbers and bone formation, but in contrast to HRHF Untreated and HRHF-IgG rats, showed higher trabecular bone volume, and reduced osteoclast numbers and serum CTX-1 levels (and statistically similar to Control levels). Conclusions: HRHF loading increased bone formation in each task group, yet blocking CCN2 dampened trabecular bone catabolism by reducing osteoclast numbers and activity.
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Project description:Aging is associated with a reduction in osteoblast life span and the volume of bone formed by each basic multicellular unit. Each time bone is resorbed, less is deposited producing microstructural deterioration. Aging is also associated with insulin resistance and hyperglycemia, either of which may cause, or be the result of, a decline in undercarboxylated osteocalcin (ucOC), a protein produced by osteoblasts that increases insulin sensitivity. We examined whether glucose-loading reduces bone remodeling and ucOC in vivo and osteoblast function in vitro, and so compromises bone formation. We administered an oral glucose tolerance test (OGTT) to 18 pre and postmenopausal, nondiabetic women at rest and following exercise and measured serum levels of bone remodeling markers (BRMs) and ucOC. We also assessed whether increasing glucose concentrations with or without insulin reduced survival and activity of cultured human osteoblasts. Glucose-loading at rest and following exercise reduced BRMs in pre and postmenopausal women and reduced ucOC in postmenopausal women. Higher glucose correlated negatively, whereas insulin correlated positively, with baseline BRMs and ucOC. The increase in serum glucose following resting OGTT was associated with the reduction in bone formation markers. D-glucose (>10 mmol L(-1)) increased osteoblast apoptosis, reduced cell activity and osteocalcin expression compared with 5 mmol L(-1). Insulin had a protective effect on these parameters. Collagen expression in vitro was not affected in this time course. In conclusion, glucose exposure reduces BRMs in women and exercise failed to attenuate this suppression effect. The suppressive effect of glucose on BRMs may be due to impaired osteoblast work and longevity. Whether glucose influences material composition and microstructure remains to be determined.
| S-EPMC4758933 | biostudies-other
Project description:Fibrosis may be a key factor in sensorimotor dysfunction in patients with chronic overuse-induced musculoskeletal disorders. Using a clinically relevant rodent model, in which performance of a high demand handle-pulling task induces tissue fibrosis and sensorimotor declines, we pharmacologically blocked cellular communication network factor 2 (CCN2; connective tissue growth factor) with the goal of reducing the progression of these changes. Young adult, female Sprague-Dawley rats were shaped to learn to pull at high force levels (10 min/day, 5 weeks), before performing a high repetition high force (HRHF) task for 3 weeks (2 h/day, 3 days/week). HRHF rats were untreated, or treated in task weeks 2 and 3 with a monoclonal antibody that blocks CCN2 (FG-3019), or a control immunoglobulin G (IgG). Control rats were untreated or received FG-3019, IgG, or vehicle (saline) injections. Mean task reach rate and grasp force were higher in 3-week HRHF + FG-3019 rats, compared with untreated HRHF rats. Grip strength declined while forepaw mechanical sensitivity increased in untreated HRHF rats, compared with controls; changes improved by FG-3019 treatment. The HRHF task increased collagen in multiple tissues (flexor digitorum muscles, nerves, and forepaw dermis), which was reduced with FG-3019 treatment. FG-3019 treatment also reduced HRHF-induced increases in CCN2 and transforming growth factor β in muscles. In tendons, FG-3019 reduced HRHF-induced increases in CCN2, epitendon thickening, and cell proliferation. Our findings indicate that CCN2 is critical to the progression of chronic overuse-induced multi-tissue fibrosis and functional declines. FG-3019 treatment may be a novel therapeutic strategy for overuse-induced musculoskeletal disorders. © 2019 The Authors. Journal of Orthopaedic Research® published by Wiley Periodicals, Inc. on behalf of Orthopaedic Research Society. J Orthop Res 37:2004-2018, 2019.
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Project description:Reducing proteinuria is a crucial approach in preventing kidney function loss. Previous preclinical studies indicated that caloric restriction (CR) imposed at a young age protects against age-related proteinuria. However, these studies have not explored CR in established renal disease. Therefore, this study aimed to investigate the impact of CR on established proteinuria. Rats, aged 12 ± 2 weeks, were administered 2.1 mg/kg of Adriamycin. Six weeks after injection, protein excretion was measured, and a [13 N]ammonia positron emission tomography (PET) scan was conducted to assess kidney perfusion. After 7 weeks rats were divided into four groups: ad libitum (AL) and CR groups fed either a 12% or a 20% protein diet. All groups were treated for 12 weeks. Blood pressure was measured and a second PET scan was acquired at the end of the study. The animals subjected to CR exhibited a 20.3% decrease in protein excretion (p = 0.003) compared to those in the AL groups. Additionally, blood pressure in the CR group was 21.2% lower (p < 0.001) than in the AL groups. While kidney function declined over time in all groups, the 20% CR group demonstrated the smallest decline. Thus CR effectively reduces urinary protein excretion and lowers blood pressure in rats with established proteinuria.
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Project description:Metastatic prostate cancer in the bone induces bone-forming lesions that contribute to progression and therapy resistance. Prostate cancer-induced bone formation originates from endothelial cells (EC) that have undergone endothelial-to-osteoblast (EC-to-OSB) transition in response to tumor-secreted BMP4. Current strategies targeting prostate cancer-induced bone formation are lacking. Here, we show that activation of retinoic acid receptor (RAR) inhibits EC-to-OSB transition and reduces prostate cancer-induced bone formation. Treatment with palovarotene, an RARγ agonist being tested for heterotopic ossification in fibrodysplasia ossificans progressiva, inhibited EC-to-OSB transition and osteoblast mineralization in vitro and decreased tumor-induced bone formation and tumor growth in several osteogenic prostate cancer models, and similar effects were observed with the pan-RAR agonist all-trans-retinoic acid (ATRA). Knockdown of RARα, β, or γ isoforms in ECs blocked BMP4-induced EC-to-OSB transition and osteoblast mineralization, indicating a role for all three isoforms in prostate cancer-induced bone formation. Furthermore, treatment with palovarotene or ATRA reduced plasma Tenascin C, a factor secreted from EC-OSB cells, which may be used to monitor treatment response. Mechanistically, BMP4-activated pSmad1 formed a complex with RAR in the nucleus of ECs to activate EC-to-OSB transition. RAR activation by palovarotene or ATRA caused pSmad1 degradation by recruiting the E3-ubiquitin ligase Smad ubiquitination regulatory factor1 (Smurf1) to the nuclear pSmad1/RARγ complex, thus blocking EC-to-OSB transition. Collectively, these findings suggest that palovarotene can be repurposed to target prostate cancer-induced bone formation to improve clinical outcomes for patients with bone metastasis.SignificanceThis study provides mechanistic insights into how RAR agonists suppress prostate cancer-induced bone formation and offers a rationale for developing RAR agonists for prostate cancer bone metastasis therapy. See related commentary by Bhowmick and Bhowmick, p. 2975.
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Project description:In this study, we investigated the effect of CCN2 (cellular communication network factor 2), previously termed connective tissue growth factor, deposited in bone matrix on osteoclastogenesis and osteoblast differentiation. To mimic the bone matrix environment, osteocytic MLO-Y4 cells had been embedded in collagen-gel with recombinant CCN2 (rCCN2), and mouse macrophage-like RAW264.7 cells were inoculated on the gel and treated with receptor activator of NF-κB ligand (RANKL). NFATc1 and cathepsin K (CTSK) productions were more increased in the combination of RAW264.7 and MLO-Y4 cells treated with rCCN2 than the combination without rCCN2. Next, we isolated an osteocyte-enriched population of cells and osteoclast progenitor cells from wild type and tamoxifen-inducible Ccn2-deficient (KO) mice and performed similar analysis. NFATc1 and CTSK productions were decreased in the KO osteocyte-enriched population at 6 months after the tamoxifen injection, regardless of the origin of the osteoclast progenitor cells. Interestingly, CTSK production was rather increased in KO osteocytes at 1 year after the injection. Finally, the combination of osteoblastic MC3T3-E1 and MLO-Y4 cells in rCCN2-containing bone matrix revealed the up-regulation of osteoblastic marker genes. These findings suggest that CCN2 supplied by osteocytes regulates both osteoclastogenesis and osteoblast differentiation.
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Project description:This study investigates the cardioprotective effects of intense caloric restriction (ICR) from birth in ovariectomized rats, a model of estrogen deficiency mimicking menopause. Our findings demonstrate that ICR significantly improved both basal and post-ischemic cardiac function, even in the absence of estrogens. The restricted animals exhibited enhanced cardiac contractility and relaxation, particularly after ischemia/reperfusion (I/R) injury, with superior functional recovery compared to control groups. Notably, ICR reduced key cardiometabolic risk factors, including blood pressure, heart rate, and adiposity, while improving glucose tolerance and insulin sensitivity. Additionally, while mitochondrial biogenesis remained unaffected, ICR preserved mitochondrial integrity by reducing the number of damaged mitochondria. This was linked to a reduction in oxidative stress, as evidenced by lower reactive oxygen species (ROS) production in the hearts of restricted animals. These results suggest that ICR offers a protective effect against cardiovascular dysfunction induced by estrogen depletion, potentially through enhanced antioxidant defenses and mitochondrial protection.
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Project description:Intense thunderstorms produce rapid cloud updrafts and may be associated with a range of destructive weather events. An important ingredient in measures of the potential for intense thunderstorms is the convective available potential energy (CAPE). Climate models project increases in summertime mean CAPE in the tropics and subtropics in response to global warming, but the physical mechanisms responsible for such increases and the implications for future thunderstorm activity remain uncertain. Here, we show that high percentiles of the CAPE distribution (CAPE extremes) also increase robustly with warming across the tropics and subtropics in an ensemble of state-of-the-art climate models, implying strong increases in the frequency of occurrence of environments conducive to intense thunderstorms in future climate projections. The increase in CAPE extremes is consistent with a recently proposed theoretical model in which CAPE depends on the influence of convective entrainment on the tropospheric lapse rate, and we demonstrate the importance of this influence for simulated CAPE extremes using a climate model in which the convective entrainment rate is varied. We further show that the theoretical model is able to account for the climatological relationship between CAPE and a measure of lower-tropospheric humidity in simulations and in observations. Our results provide a physical basis on which to understand projected future increases in intense thunderstorm potential, and they suggest that an important mechanism that contributes to such increases may be present in Earth's atmosphere.
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