Protective effects of Phlomis umbrosa extract on a monosodium iodoacetate-induced osteoarthritis model and prediction of molecular mechanisms using transcriptomics.

BACKGROUND: Phlomis umbrosa Turczaninow root has been traditionally used to treat fractures, rheumatoid arthritis, and arthralgia. However, the effects and mechanisms of P. umbrosa on osteoarthritis (OA) remain poorly understood and a functional genomic approach has not been investigated. AIM: The purpose of this study was to investigate the effects and mechanisms of P. umbrosa extract (PUE) on OA using transcriptomic analysis. METHODS: We performed joint diameter measurements, micro computed tomography, and histopathological analysis of monosodium iodoacetate (MIA)-induced OA rats treated with PUE (200 mg/kg) for 3 weeks. Gene expression profiling in articular cartilage tissue was then performed using RNA sequencing (RNA-seq) followed by signaling pathway analysis of regulatory genes. RESULTS: PUE treatment improved OA based on decreased joint diameter, increased joint morphological parameters, and histopathological features. Many genes involved in multiple signal transduction pathway and collagen activation in OA were differentially regulated by PUE. These included genes related to Wnt/ß-catenin, OA pathway, and sonic hedgehog signaling activity. Furthermore, PUE treatment downregulated cartilage damage factors (MMP-9, MMP-13, ADAMTs4, and ADMATs5) and upregulated chondrogenesis (COL2A1 and SOX-9) by regulating the transcription factors SOX-9, Ctnnb1, and Epas1. CONCLUSION: Based on the results of gene expression profiling, this study highlighted the molecular mechanisms underlying the effects of PUE in MIA-induced OA rats. The findings provide novel insight into the mechanisms by which PUE treatment-induced gene expression changes may influence OA disease progression. Taken together, the results suggest that PUE may be used as a source of therapeutic agents for OA.

In vivo protective effects of upper zone of growth plate and cartilage matrix associated protein against cartilage degeneration in a monosodium iodoacetate induced osteoarthritis model.

Osteoarthritis (OA) is a degenerative disease affecting the majority of over 65 year old people and characterized by cartilage degeneration, subchondral abnormal changes, and inflammation. Despite the enormous socioeconomic burden caused by OA, currently, there is no effective therapy against it. Upper zone of growth plate and cartilage matrix associated protein (UCMA) is a vitamin K dependent protein and has a critical role in pathophysiological conditions associated with bone and cartilage. However, there is no research on the protective role of intra-articular UCMA treatment in OA pathogenesis. Therefore, we aimed to investigate the potential therapeutic role of UCMA in an in vivo model of OA. We report for the first time that intra-articular UCMA injection ameliorated cartilage degeneration in a monosodium iodoacetate induced OA rat model. Furthermore, the OA-induced activation of nuclear factor kappa B and bone morphogenetic protein 2 signals was attenuated by UCMA. Our results indicated that UCMA decreased cartilage oligomeric matrix protein levels but did not affect interleukin 6, total antioxidant status, and total oxidant status levels in the serum. In conclusion, UCMA exhibited a therapeutic potential in the treatment of OA. This protective effect of UCMA is possibly achieved by reducing the aggrecanase activity and the production of inflammatory cytokines.

Effect of Alpinia oxyphylla extract in vitro and in a monosodium iodoacetate-induced osteoarthritis rat model.

BACKGROUND: Osteoarthritis (OA) affects the articular cartilage and subchondral bone of synovial joints and induces proinflammatory and anti-inflammatory pathway dysregulation, leading to pain. This study evaluated the anti-inflammatory and antiosteoarthritis effects of Alpinia oxyphylla extract (AOE) in vitro and in vivo. METHODS: The anti-inflammatory effect of AOE was evaluated in vitro in lipopolysaccharide (LPS)-treated RAW264.7 cells. The antiosteoarthritis effect of AOE was investigated in a monosodium iodoacetate (MIA)-induced rat model of OA. Rats were orally administered AOE (150 mg/kg or 300 mg/kg) or the positive control drug indomethacin (1 mg/kg) 3 days before MIA injection and once daily for 21 days thereafter. RESULTS: AOE significantly decreased the production of nitric oxide (NO, 68.2%), prostaglandin E2 (PGE2, 92.8%), interleukin-1ß (IL-1ß, 77.2%), interleukin-6 (IL-6, 39.9%), and tumor necrosis factor-alpha (TNF-α, 20.7%) and the activation of extracellular signal-regulated kinase (ERK), Jun N-terminal kinase (JNK) and p38 mitogen-activated protein kinase (MAPK) in LPS-treated RAW264.7 cells at a dose of 100 µg/ml. In addition, AOE attenuated joint pain, suppressed proinflammatory cytokine and mediator production and inhibited cartilage degradation in the MIA-induced rat OA model. CONCLUSION: These results demonstrate that AOE exerts potent anti-inflammatory effects and may be a useful therapeutic candidate against OA.

Mori Folium water extract alleviates articular cartilage damages and inflammatory responses in monosodium iodoacetate­induced osteoarthritis rats.

Mori folium, the leaf of Morus alba L. (Moraceae), has been widely used in traditional medicine for the treatment of various diseases. It has been recently reported that Mori folium possesses potential chondroprotective effects in interleukin (IL)­1ß­stimulated human chondrocytes; however, its protective and therapeutic potential against osteoarthritis (OA) in an animal model remains unclear. In this study, as part of an ongoing screening program to evaluate the anti­osteoarthritic potential of Mori folium, the protective effects of a water extract of Mori folium (MF) on cartilage degradation and inflammatory responses in a monosodium iodoacetate (MIA)­induced OA rat model were evaluated. The results demonstrated that administration of MF had a tendency to attenuate the damage to articular cartilage induced by MIA, as determined by knee joint swelling and the histological grade of OA. The elevated levels of matrix metalloproteinases­13 and two bio­markers for the diagnosis and progression of OA, such as the cartilage oligomeric matrix protein and C­telopeptide of type II collagen, were markedly ameliorated by MF administration in MIA­induced OA rats. In addition, MF significantly suppressed the production of pro­inflammatory cytokines, including IL­1ß, IL­6 and tumor necrosis factor­α. MF also effectively inhibited the expression of inducible nitric oxide (NO) synthase and cyclooxygenase­2, thus inhibiting the release of NO and prostaglandin E2. Although further work is required to fully understand the critical role and clinical usefulness, these findings indicate that MF may be a potential therapeutic option for the treatment of OA.

Effect of alcoholic extract of Entada pursaetha DC on monosodium iodoacetate-induced osteoarthritis pain in rats.

BACKGROUND & OBJECTIVES: Osteoarthritis (OA) is a degenerative disease characterized by joint pain and progressive loss of articular cartilage. Entada pursaetha has been traditionally used in the treatment of inflammatory disease, liver ailment, etc. In this study we investigated suppressive effect of ethanolic extract of E. pursaetha (EPE) on monosodium iodoacetate (MIA)-induced osteoarthritis pain and disease progression by histopathological changes in joints in a rat model. METHODS: OA was induced in right knee of rat by intra-articular injection of 3 mg of MIA and characterized by pathological progression of disease and pain of affected joint. Spontaneous movements, mechanical, thermal and cold sensitivity were monitored at days 0 (before drug and MIA injection), 7, 14 and 21 of MIA administration. EPE (30, 100 and 300 mg/kg), vehicle or etoricoxib (10 mg/ kg; reference drug) were administered daily for 21 days by oral route. RESULTS: EPE at various doses significantly reduced mechanical, heat, cold hyperalgesia and increased the horizontal and vertical movements in intra-articular MIA injected rats. EPE prevented the damage to cartilage structure and reduced the cellular abnormalities. Articular cartilage of rats treated with EPE at 300 mg/kg group was almost normal with well-developed smooth surface and chondrocytes were distributed individually or arranged in column. INTERPRETATION & CONCLUSIONS: The present findings showed that the EPE was not only able to mitigate pain and hyperalgesia but also inhibited MIA-induced cartilage degeneration in vivo. EPE may have the potential to become therapeutic modality in the treatment of osteoarthritis. However, further studies need to be done to confirm these findings in other models and clinical trials.

The inhibition of subchondral bone lesions significantly reversed the weight-bearing deficit and the overexpression of CGRP in DRG neurons, GFAP and Iba-1 in the spinal dorsal horn in the monosodium iodoacetate induced model of osteoarthritis pain.

BACKGROUND: Chronic pain is the most prominent and disabling symptom of osteoarthritis (OA). Clinical data suggest that subchondral bone lesions contribute to the occurrence of joint pain. The present study investigated the effect of the inhibition of subchondral bone lesions on joint pain. METHODS: Osteoarthritic pain was induced by an injection of monosodium iodoacetate (MIA) into the rat knee joint. Zoledronic acid (ZOL), a third generation of bisphosphonate, was used to inhibit subchondral bone lesions. Joint histomorphology was evaluated using X-ray micro computed tomography scanning and hematoxylin-eosin staining. The activity of osteoclast in subchondral bone was evaluated using tartrate-resistant acid phosphatase staining. Joint pain was evaluated using weight-bearing asymmetry, the expression of calcitonin gene-related peptide (CGRP) in the dorsal root ganglion (DRG), and spinal glial activation status using glial fibrillary acidic protein (GFAP) and ionized calcium binding adaptor molecule-1 (Iba-1) immunofluorescence. Afferent neurons in the DRGs that innervated the joints were identified using retrograde fluorogold labeling. RESULTS: MIA injections induced significant histomorphological alterations and joint pain. The inhibition of subchondral bone lesions by ZOL significantly reduced the MIA-induced weight-bearing deficit and overexpression of CGRP in DRG neurons, GFAP and Iba-1 in the spinal dorsal horn at 3 and 6 weeks after MIA injection; however, joint swelling and synovial reaction were unaffected. CONCLUSIONS: The inhibition of subchondral bone lesions alleviated joint pain. Subchondral bone lesions should be a key target in the management of osteoarthritic joint pain.

Monosodium iodoacetate-induced osteoarthritis produces pain-depressed wheel running in rats: implications for preclinical behavioral assessment of chronic pain.

Pain stimulates some behaviors (e.g., withdrawal responses) and depresses other behaviors (e.g., feeding and locomotion). We are developing methods for testing candidate analgesics using measurements of pain-depressed behaviors. Such assays may model important aspects of clinical pain and complement traditional procedures that measure pain-stimulated behaviors. The present study characterized the effects of a chronic pain manipulation (monosodium iodoacetate (MIA)-induced osteoarthritis) on wheel running in rats. Rats had 24 h voluntary access to running wheels. Duration of running wheel acquisition was manipulated such that rats had either 21 or 7 days of running wheel access prior to MIA administration. Wheel running was monitored for an additional 21 days following MIA administration. MIA produced concentration- and acquisition length-dependent decreases in wheel running. Parallel experiments demonstrated that MIA produced concentration-dependent tactile allodynia and shifts in hind limb weight bearing. MIA was differentially potent across assays with a potency rank: weight-bearing≥von Frey>running wheel. MIA produced greater depression of wheel running in rats with relatively high baseline running rates compared to rats with relatively low baseline running rates. The differential potency of MIA across assays and apparent rate-dependent effects in running wheels may impact our traditional interpretations of preclinical nociceptive and antinociceptive testing.

The effect of nordihydroguaiaretic acid on iodoacetate-induced toxicity in cultured neurons.

Nordihydroguaiaretic acid (NDGA) is present in high concentrations in the desert shrub Creosote bush, Larrea tridentate. This plant has been used in traditional medicine because of its beneficial effects related, at least in part, to its antioxidant properties. Taking into account some evidence about neuroprotective effects elicited by NDGA, we evaluated the effect of this compound on the neurotoxicity induced by iodoacetate (IAA), an inhibitor of glyceraldehyde-3-phosphate dehydrogenase (GAPDH), on cerebellar granule neurons. In addition, as reactive oxygen species play an important role in IAA-induced cytotoxicity, we also studied the enzymatic antioxidant system in IAA-treated cells. We found that IAA caused a dose-dependent decrease in cell viability of cultured neurons with an IC(50) of 18.4 microM and induced increased activity of catalase, glutathione peroxidase, and glutathione-S-transferase. Moreover, NDGA attenuated the toxicity induced by 18.4, 25, and 30 microM of IAA without abolishing the inhibitory effect of IAA on GAPDH activity. Furthermore, NDGA could prevent the inhibitory effect of IAA on aconitase activity, a marker of oxidative stress, suggesting that the protective effect of NDGA on IAA neurotoxicity was associated with the prevention of oxidative stress.

Vasoactive intestinal peptide (VIP) is a modulator of joint pain in a rat model of osteoarthritis.

Osteoarthritis (OA) is a debilitating disease in which primarily weight-bearing joints undergo progressive degeneration. Despite the widespread prevalence of OA in the adult population, very little is known about the factors responsible for the generation and maintenance of OA pain. Vasoactive intestinal peptide (VIP) was identified in the synovial fluid of arthritis patients nearly 20 years ago and the aim of this study was to examine whether VIP could be involved in the generation of OA pain. Hindlimb weight bearing was used as a measure of joint pain, while von Frey hair algesiometry applied to the plantar surface of the ipsilateral hindpaw tested for secondary mechanical hyperalgesia. Intra-articular injection of VIP into normal rat knee joints caused a significant shift in weight bearing in favour of the contralateral non-injected hindlimb as well as causing a reduction in ipsilateral paw withdrawal threshold. These pain responses were blocked by co-administration of the VPAC receptor antagonist VIP6-28. Induction of OA by intra-articular sodium monoiodoacetate injection resulted in a reduction in weight bearing on the affected leg, but no evidence of secondary hyperalgesia in the paw. Treatment of OA knees with a single injection of VIP6-28 diminished hindlimb incapacitance while increasing paw withdrawal threshold. This study showed for the first time that peripheral application of VIP causes increased knee joint allodynia and secondary hyperalgesia. Furthermore, antagonists that inhibit VIP activity may prove beneficial in the alleviation of OA pain.

In vivo micro computed tomography of subchondral bone in the rat after intra-articular administration of monosodium iodoacetate.

Osteoarthritis (OA) is a degenerative disease that is characterized by joint discomfort, loss of articular cartilage, and changes to the subchondral bone. Studies to elucidate the pathophysiology of OA have been hampered by the lack of a rapid, reproducible animal model that mimics the structural changes associated with the disease. A single intra-articular injection of mono-iodoacetate (MIA), an inhibitor of glycolysis, into the femorotibial joint of rodents promotes loss of articular cartilage similar to that noted in human OA. The purpose of the present study was to determine whether in vivo three-dimensional micro computed tomography (microCT) was of use for detecting progressive changes over time to the subchondral bone (femorotibial joint) of Wistar rats treated with a single intra-articular injection of MIA. MIA-treated right knee joints and left contralateral control knee joints were imaged in vivo at 0, 1, 7, 14, 28, and 56 days postinjection by using microCT. Analysis of 50- and 100- micro m resolution images demonstrated that changes to the subchondral bone, as determined by visual and bone mineral density analysis, are apparent by day 14 post-MIA. By day 28, there were marked changes to lateral aspect of the medial tibial plateaus of the subchondral bone in MIA-treated joints. These changes were progressive through day 56. It was concluded that intra-articular injection of MIA induces progressive changes to subchondral bone that can be assessed using in vivo microCT imaging. In light of these data, in vivo microCT imaging represents a valuable tool for investigating bone remolding and has the potential to be used for routine, high-throughput analysis and screening of investigation therapeutics.