Alendronate-functionalized porous nano-crystalsomes mitigate osteolysis and consequent inhibition of tumor growth in a tibia-induced metastasis model.

Bone is one of the most prevalent sites of metastases in various epithelial malignancies, including breast cancer and this metastasis to bone often leads to severe skeletal complications in women due to its osteolytic nature. To address this, we devised a novel drug delivery approach using an Alendronate (ALN) functionalized self-assembled porous crystalsomes for concurrent targeting of Oleanolic acid (OA) and ALN (ALN + OA@NCs) to bone metastasis. Initially, the conjugation of both PEG-OA and OA-PEG-ALN with ALN and OA was achieved, and this conjugation was then self-assembled into porous crystalsomes (ALN + OA@NCs) by nanoemulsion crystallization. The reconstruction of a 3D single particle using transmission electron microscopy ensured the crystalline porous structure of ALN + OA@NCs, was well aligned with characteristic nanoparticle attributes including size distribution, polydispersity, and zeta potential. Further, ALN + OA@NCs showed enhanced efficacy in comparison to OA@NCs suggesting the cytotoxic roles of ALN towards cancer cells, followed by augmentation ROS generation (40.81%), mitochondrial membrane depolarization (57.20%), and induction of apoptosis (40.43%). We found that ALN + OA@NCs facilitated inhibiting osteoclastogenesis and bone resorption followed by inhibited osteolysis. In vivo activity of ALN + OA@NCs in the 4 T1 cell-induced tibia model rendered a reduced bone loss in the treated mice followed by restoring bone morphometric markers which were further corroborated bone-targeting effects of ALN + OA@NCs to reduce RANKL-stimulated osteoclastogenesis. Further, In vivo intravenous pharmacokinetics showed the improved therapeutic profile of the ALN + OA@NCs in comparison to the free drug, prolonging the levels of the drug in the systemic compartment by reducing the clearance culminating the higher accumulation at the tumor site. Our finding proposed that ALN + OA@NCs can effectively target and treat breast cancer metastasis to bone and its associated complications.

Linoleic acid blunts early osteoblast differentiation and impairs oxidative phosphorylation in vitro.

BACKGROUND: Linoleic acid (LNA), an essential polyunsaturated fatty acid (PUFA), plays a crucial role in cellular functions. However, excessive intake of LNA, characteristic of Western diets, can have detrimental effects on cells and organs. Human observational studies have shown an inverse relationship between plasma LNA concentrations and bone mineral density. The mechanism by which LNA impairs the skeleton is unclear, and there is a paucity of research on the effects of LNA on bone-forming osteoblasts. METHODS: The effect of LNA on osteoblast differentiation, cellular bioenergetics, and production of oxidized PUFA metabolites in vitro, was studied using primary mouse bone marrow stromal cells (BMSC) and MC3T3-E1 osteoblast precursors. RESULTS: LNA treatment decreased alkaline phosphatase activity, an early marker of osteoblast differentiation, but had no effect on committed osteoblasts or on mineralization by differentiated osteoblasts. LNA suppressed osteoblast commitment by blunting the expression of Runx2 and Osterix, key transcription factors involved in osteoblast differentiation, and other key osteoblast-related factors involved in bone formation. LNA treatment was associated with increased production of oxidized LNA- and arachidonic acid-derived metabolites and blunted oxidative phosphorylation, resulting in decreased ATP production. CONCLUSION: Our results show that LNA inhibited early differentiation of osteoblasts and this inhibitory effect was associated with increased production of oxidized PUFA metabolites that likely impaired energy production via oxidative phosphorylation.

A Novel Blockchain and Internet of Things-Based Food Traceability System for Smart Cities.

Rapid urbanization has recently caused serious problems for cities all around the world. Smart cities have drawn much interest from researchers in the present research paradigm to manage the expanding urban population. Frameworks for smart cities are planned and implemented using platforms based on blockchain and the Internet of Things (BIOT). Smart cities may use the BIoT platform to provide improved transportation, food traceability, and healthcare services. Food safety is one of the sectors where less research has been done than the others. The importance of food safety is now more widely recognized, making it essential to improve the traceability and transparency of the food supply chain. In this paper, a novel BIOT-based layered framework using EOSIO has been proposed for effective food traceability. The proposed system first identifies the suitable traceability units to provide better transparency and traceability and then defines and implements a layered architecture using Ethereum and EOSIO blockchain platforms. The performance of the proposed EOSIO-based model is evaluated using the practicality of the consensus algorithm, block production rate, throughput, and block confirmation time. The proposed traceability system attains a block production rate of 0.5 s and a block confirmation time of 1 s, which is much lower than the Ethereum-based traceability system. Hence, from the experimental evidence, the superiority of the proposed EOSIO-based food traceability can be observed.

Nanoparticles of naturally occurring PPAR-γ inhibitor betulinic acid ameliorates bone marrow adiposity and pathological bone loss in ovariectomized rats via Wnt/ß-catenin pathway.

AIMS: Postmenopausal osteoporosis is one of the world's biggest yet unnoticed health issues. After ovariectomy, declined estrogen level significantly contributes to the elevation of bone marrow adiposity and bone loss leading to osteoporosis. Therapeutics to prevent osteoporosis addressing various aspects are now in short supply. In this study we made an approach to synthesize nanoparticles of naturally occurring PPAR-γ inhibitor, betulinic acid (BA/NPs) and tested the same in altered bone metabolisms developed after ovariectomy. MAIN METHODS: The osteogenic efficacy of BA/NPs was established in human and rat primary osteoblast cells using qRT-PCR and immunoblot analysis. Furthermore, lineage allocations of multipotent bone marrow stromal cells were evaluated. Various aspects of altered bone metabolism after ovariectomy such as bone marrow adiposity and pathological bone loss were evaluated using µCT and histological assessments. KEY FINDINGS: BA/NPs exert potential osteogenic efficacy by modulating key osteogenic markers such as RUNX2 and BMP2. Mechanistically BA/NPs regulate osteoblastogenesis through Wnt/ß-catenin signaling. Further, BA/NPs showed the potential to inhibit the differentiation of multipotent BMSCs towards adipogenesis while favouring the osteogenic lineage via Wnt/ß-catenin pathway. In the in vivo study, increased bone marrow adiposity was reduced in ovariectomized rats after BA/NPs treatment as assessed by histology and µCT analysis. Loss of bone mineral density as a hallmark of pathological bone loss was also abrogated by BA/NPs. Furthermore, increased obesity after OVX was also prevented in BA/NPs treated animals. SIGNIFICANCE: Our findings imply that BA/NPs could be used further as a viable drug lead to counteract various pathophysiological challenges after menopause.

A novel BMP2 secretagogue ameliorates glucocorticoid induced oxidative stress in osteoblasts by activating NRF2 dependent survival while promoting Wnt/ß-catenin mediated osteogenesis.

In our previous study, a novel BMP2 secretagogue was synthesized belonging to a class of galloyl conjugates of flavanones, with remarkable osteogenic potential that promoted bone regeneration. We aimed to establish the protective effect of our compound against bone loss that co-exists with excess Glucocorticoid (GC) therapy. GC therapy induces osteoblast damage leading to apoptosis by increasing reactive oxygen species (ROS). Our results delineate that compound 5e (a BMP2 secretagogue) activates NRF2 signalling to counter the disturbed cellular redox homeostasis and escalate osteoblast survival as assessed by Western blot and immunocytochemistry. Depletion of NRF2 by siRNA blocked activation of the NRF2/HO-1 pathway, magnified oxidative stress, increased apoptosis and abrogated the protective effects of compound 5e. 5e, on the other hand, increased ALP, mineralization activity, and promoted osteoblast differentiation by activating WNT/ß-catenin signalling in BMP2 dependent manner, validated by Western blot of WNT3A, SOST, GSK3-ß and ß-catenin nuclear translocation. Treatment of 5e in presence of BMP inhibitor noggin attenuated the osteogenic efficacy and minimized Wnt//ß-catenin signalling in presence of dexamethasone. Our compound prevents GC challenged trabecular and cortical bone loss assessed by micro-CT and promotes bone formation and osteocyte survival determined by calcein labelling and TUNEL assay in GC treated animals. The osteogenic potential of the compound was authenticated by bone turnover markers. On a concluding note, compounds with BMP upregulation can be potential therapeutics for the prevention and treatment of glucocorticoid-induced osteoporosis.

Benzofuran pyran hybrid prevents glucocorticoid induced osteoporosis in mice via modulation of canonical Wnt/ß-catenin signaling.

Glucocorticoid induced osteoporosis (GIOP) is the second most leading cause of osteoporosis. We have identified a compound, a benzofuran pyran hybrid compound 4e that has osteogenic potential and we wanted to assess its efficacy in GIOP in male mice. We assessed the effect of dexamethasone and compound 4e on primary osteoblasts using various cell based and immunofluorescence assays. For in vivo studies we administered methylprednisolone and compound 4e as a prophylactic measure in male Balb/c mice for 28 days and then evaluated the effect on bone microarchitecture by microCT, bone formation by histology along with clinically relevant bone markers. Compound 4e preserved osteoblast differentiation as evident by higher ALP positive cells and mineralization in compound treated groups. Compound 4e also increased the expression of osteogenic genes. This compound guarded ß-catenin expression both in vitro and in vivo as confirmed by western blot and immunofluorescence assays. This led to the preservation of bone microarchitecture and cortical thickness at 2.5 mg kg-1 and 5 mg kg-1 doses. Further compound 4e enhanced bone formation rate and regulated osteocyte death. The osteogenic potential of compound 4e was reflected by an increased level of serum marker osteocalcin and decreased levels of SOST and CTX-I. Overall, Compound 4e is able to overcome the catabolic effect of dexamethasone on bone by targeting the canonical WNT/ß-catenin signaling as evidenced by both in vitro and in vivo studies.

Caviunin glycoside (CAFG) from Dalbergia sissoo attenuates osteoarthritis by modulating chondrogenic and matrix regulating proteins.

ETHNOPHARMACOLOGICAL RELEVANCE: Dalbergia sissoo DC. (Indian rosewood or Sheesham) is a traditional medicinal plant, reported since time immemorial for its analgesic, anti-nociceptive, anti-inflammatory, and immuno-modulatory properties. D. sissoo DC (DS). is being used traditionally to cure joint inflammation and joint pain. AIM: To study the potential of DS leaves and its derived novel compound CAFG to treat the clinical symptoms of osteoarthritis (OA) and its underlying mechanism. METHODS: The chemical profile of DS extract (DSE) with isoflavonoids and isoflvaonoid glycosides from the DS was established by UHPLC-PDA and UHPLC-MS/MS. Monosodium iodoacetate (MIA) was injected into the knee joint to develop the OA model in rats. DSE was given orally for 28 days daily at 250 and 500 mg.kg-1day-1. For in-vitro experiments, chondrocytes isolated from joint articular cartilage were negatively induced with interleukin-1ß (IL-1ß) and CAFG was given to the cells as a co-treatment. RESULTS: Chondrocytes undergo apoptosis following inflammation and proteoglycan synthesis affected in MIA injected knees. DSE administration prevented these effects as assessed by H&E and Toluidine blue staining. Micro-CT analysis showed that subchondral bone loss was restored. DSE decreased elevated serum levels of cartilage-bone degradation (CTX-I, CTX-II, and COMP), inflammation markers IL-1ß, and matrix-degrading MMP-3 and 13. The effects of IL-1ß on gene expression of chondrocytes were reversed by CAFG treatment at 1 µM. CONCLUSION: Data showed that DSE protected joint cartilage and deterioration in subchondral bone in vivo while in in-vitro, its active ingredient CAFG prevented interleukin-1ß induced effects and inhibited OA. This finding suggest that DSE and CAFG could be used as a possible therapeutic to treat osteoarthritis.

Synthesis and Evaluation of Galloyl Conjugates of Flavanones as BMP-2 Upregulators with Promising Bone Anabolic and Fracture Healing Properties.

The molecular hybridization concept led us to design a series of galloyl conjugates of flavanones that have potent osteoblast differentiation ability in vitro and promote bone formation in vivo. An array of in vitro studies, especially gene expression of osteogenic markers, evinced compound 5e as the most potent bone anabolic agent, found to be active at 1 pM, which was then further assessed for its osteogenic potential in vivo. From in vivo studies on rat calvaria and a fracture defect model, we inferred that compound 5e, at an oral dose of 5 mg/(kg day), increased the expression of osteogenic genes (RUNX2, BMP-2, Col1, and OCN) and the bone formation rate and significantly promoted bone regeneration at the fracture site, as evidenced by the increased bone volume/tissue fraction compared with vehicle-treated rats. Furthermore, structure-activity relationship studies and pharmacokinetic studies suggest 5e as a potential bone anabolic lead for future osteoporosis drug development.

Inhibition of cartilage degeneration and subchondral bone deterioration by Spinacia oleracea in human mimic of ACLT-induced osteoarthritis.

Osteoarthritis (OA) is an aging disorder characterized by degenerated cartilage and sub-chondral bone alteration in affected knee joints. Globally, millions of people suffer from this disease. However, there is a lack of safe and promising therapeutics, making the exploration and development of leads from natural sources urgent. Accordingly, food as medicine may be the most suitable approach for the treatment of this degenerative disease. Herein, we elucidated the protective role of Spinacia oleracea extract (SOE) in an anterior cruciate ligament transection (ACLT) model of osteoarthritis as a mimic of the human condition. ACL transection was done in the tibio-femoral joints of rats. SOE was orally administered at the dosage of 125 and 250 mg kg-1 day-1 for four weeks. It was shown that the animals with SOE treatment had better joint morphology than the ACLT animals, as evident by the shiny appearance of their cartilage. Hematoxylin and safranin-o staining showed that the number of chondrocytes was significantly reduced in the OA model, which was prevented with SOE treatment. The reduction in the cartilage thickness was well observed by toluidine blue staining. The reduced stain by safranin-o and toluidine blue, indicated proteoglycan loss in the ACLT-induced osteoarthritis model. The proteoglycan content and cartilage thickness were restored in the SOE group upon treatment at an SOE dosage of 125 and 250 mg kg-1 day-1. The micro-CT parameters of subchondral bone (SCB) and cartilage degradation markers in the serum corroborated our findings of the protective effects of SOE. In summary, our study suggests that SOE has therapeutic potential, which if taken regularly as a food supplement, can have beneficial effects.

Benzofuran pyran compound rescues rat and human osteoblast from lipotoxic effect of palmitate by inhibiting lipid biosynthesis and promoting stabilization of RUNX2.

Obesity and ageing increases bone marrow fat which in turn is associated with lower bone mass. Marrow adipocytes by secreting cytokines, adipokines and free fatty acids change the bone marrow milieu and thus the number of osteoblasts. Palmitate is the common saturated fatty acid, an unavoidable ingredient we consume with food, which kindles cell apoptosis. Compound 4e is osteogenic in nature. We examine the effect of compound 4e in palmitate induced lipotoxicity in rat osteoblasts. Design of benzofuran Pyran hybrid compound (4e) was found to be effective in inhibiting palmitate induced cell apoptosis. In this study an in vitro model of palmitate was contrived. Anti-apoptotic effect of compound 4e was assessed by Annexin/PI and LDH (Lactate dehydrogenase) assay. Compound 4e also increased osteoblast differentiation and mineralization. It also increased expression of osteogenic markers (RUNX2 and BMP2), assessed by Real time PCR and immunofluorescence, which was impeded by palmitate. Acetyl Co-Carboxylase (ACC) and Fatty acid synthase (FAS), two prominent mediators of lipid biosynthesis were increased by palmitate exposure. Compound 4e modulated lipid biosynthesis by inhibiting ACC and FAS as reflected visually and after quantification of less lipid droplet formation suggesting that 4e is osteogenic and simultaneously anti-lipotoxic.

Glucocorticoid aggravates bone micro-architecture deterioration and skeletal muscle atrophy in mice fed on high-fat diet.

High fat diet (HFD) induced obesity has deleterious effect on bone micro-architecture and is associated with low-grade chronic inflammation. Exogenous glucocorticoids (GC) are used to treat inflammatory conditions but with concomitant adverse effect on musculoskeletal system. This study aims to highlight the effect of exogenous GCs on musculoskeletal system in mice fed on HFD. Adult BALB/c mice were fed either normal chow or high fat diet and were exogenously administered with GC for 10 weeks. At the end of the study, animals were autopsied and bone, muscle, serum samples were collected for micro-CT, gene expression and histological study. HFD induced obesity resulted in deterioration in bone micro-architecture predominant in trabecular region of long bones and was significantly amplified with GC administration. Approximately, 37% and 25% loss in femoral and tibial bone volume was observed in obese animals with exogenous GC. Further, deteriorating bone pathology was apparent from reduced bone mineral density (BMD) and bone strength parameter which was correlated to alteration in osteoblast and adipocytes pool of cells in bone marrow. Transcriptional analysis of osteoblast marker genes, bone morphogenetic protein 2 (BMP-2), osteocalcin (OCN) exhibited decreased formation. Moreover, similar degeneration was observed in skeletal muscle physiology with stimulation in muscle atrophy genes atrogin-1, muscle ring finger motif-1 (MuRF-1) and inflammatory markers accompanied with intra-myocellular lipid accumulation. Thus, our results showed that detrimental effect of GC on bone and skeletal muscle is aggravated with HFD, attributed to alteration in bone marrow cell population and skeletal muscle atrophy.

FGF-2 targets sclerostin in bone and myostatin in skeletal muscle to mitigate the deleterious effects of glucocorticoid on musculoskeletal degradation.

AIM: Myokines are associated with regulation of bone and muscle mass. However, limited information is available regarding the impact of myokines on glucocorticoid (GC) mediated adverse effects on the musculoskeletal system. This study investigates the role of myokine fibroblast growth factor-2 (FGF-2) in regulating GC-induced deleterious effects on bone and skeletal muscle. METHODS: Primary osteoblast cells and C2C12 myoblast cell line were treated with FGF-2 and then exposed to dexamethasone (GC). FGF-2 mediated attenuation of the inhibitory effect of GC on osteoblast and myoblast differentiation and muscle atrophy was assessed through quantitative PCR and western blot analysis. Further, FGF-2 was administered subcutaneously to dexamethasone treated mice to collect bone and skeletal muscle tissue for in vivo analysis of bone microarchitecture, mechanical strength, histomorphometry and for histological alterations in treated tissue samples. KEY FINDINGS: FGF-2 abrogated the dexamethasone induced inhibitory effect on osteoblast differentiation by modulating BMP-2 pathway and inhibiting Wnt antagonist sclerostin. Further, dexamethasone induced atrophy in C2C12 cells was mitigated by FGF-2 as evident from down regulation of atrogenes expression. FGF-2 prevented GC-induced impairment of mineral density, biomechanical strength, trabecular bone volume, cortical thickness and bone formation rate in mice. Additionally, skeletal muscle tissue from GC treated mice displayed weak myostatin immunostaining and reduced expression of atrogenes following FGF-2 treatment. SIGNIFICANCE: FGF-2 mitigated GC induced effects through inhibition of sclerostin and myostatin expression in bone and muscle respectively. Taken together, this study exhibited the role of exogenous FGF-2 in sustaining osteoblastogenesis and inhibiting muscle atrophy in presence of glucocorticoid.

MicroRNA-672-5p Identified during Weaning Reverses Osteopenia and Sarcopenia in Ovariectomized Mice.

Post-menopausal condition augments the biological aging process, characterized by multiple metabolic disorders in which bone loss is the most prevalent outcome and usually coupled with sarcopenia. Coexistence of such associated pathogenesis have much worse health outcomes, compared to individuals with osteoporosis only. Pre- and post-natal bone development demands calcium from mother to fetus during pregnancy and lactation leading to a significant maternal skeletal loss. It follows an anabolic phase around weaning during which there is a notable recovery of the maternal skeleton. Here, we have studied the therapeutic effect of microRNA-672-5p identified during weaning when it is predominantly expressed, in ovariectomized mice for both osteopenia and sarcopenia. miR-672-5p induced osteoblast differentiation and mineralization. These actions were mediated through inhibition of Smurf1 with enhanced Runx2 transcriptional activation. In vivo, miR-672-5p significantly increased osteoblastogenesis and mineralization, thus reversing bone loss caused by ovariectomy. It also improved bone-mineral density, load-bearing capacity, and bone quality. Sarcopenia was also alleviated by miR-672-5p, as we observed increased cross-sectional area and Feret's diameter of muscle fibers. We hypothesize that elevated miR-672-5p expression has therapeutic efficacy in estrogen-deficiency-induced osteopenia along with sarcopenia.

Glucose dependent miR-451a expression contributes to parathyroid hormone mediated osteoblast differentiation.

Parathyroid hormone (PTH; amino acid 1-34, known as teriparatide) has reported promoting differentiation and glucose uptake in osteoblasts. However, how PTH regulates glucose metabolism to facilitate osteoblast differentiation is not understood. Here, we report that PTH promotes glucose dependent miR-451a expression which stimulates osteoblast differentiation. In addition to glucose uptake, PTH suppresses AMPK phosphorylation via PI3K-mTOR-AKT axis thereby preventing phosphorylation and inactivation of octamer-binding transcription factor 1 (OCT-1) which has been reported to act on the promoter region of miR-451a. Modulation of AMPK activity controls miR-451a levels in differentiating osteoblasts. Moreover, pharmacological inhibition of PI3K-mTOR-AKT axis suppressed miR-451a via increased AMPK activity. We report that this glucose regulated miRNA is an anabolic target and transfection of miR-451a mimic induces osteoblast differentiation and mineralization in vitro. These actions were mediated through the suppression of Odd-skipped related 1 (Osr1) and activation of Runx2 transcription. When injected in vivo, the miR-451a mimic significantly increased osteoblastogenesis, mineralization, reversed ovariectomy induced bone loss and improved bone strength. Together, these findings suggest that enhanced osteoblast differentiation associated with bone formation in case of PTH therapy is also a consequence of elevated miR-451a levels via glucose regulation. Consequently, this miRNA has the potential to be a therapeutic target for conditions of bone loss.

Synthesis and study of benzofuran-pyran analogs as BMP-2 targeted osteogenic agents.

Twenty-four novel benzofuran-pyran derivatives were synthesized and evaluated for their anti-osteoporotic activity in primary cultures of rat calvarial osteoblasts in vitro. Among all the compounds screened for the alkaline phosphatase activity, three compounds 4e, 4j and 4k showed potent activity at picomolar concentrations in osteoblast differentiating stimulation. Additionally, these compounds were found effective in mineralization, assessed by alizarin red-S staining assay. Compounds were again validated through a series of other in vitro experiments. Moreover, molecular dynamics simulations demonstrated that both benzofuran and pyran moieties are requisite to fit into the active site of BMP-2 receptor, a key target of the osteogenic agents. The obtained results strongly convey that compound 4e is a potential bone anabolic agent among synthesized series, which can be further explored as a drug lead for treating osteoporosis.

Spinacia oleracea extract attenuates disease progression and sub-chondral bone changes in monosodium iodoacetate-induced osteoarthritis in rats.

BACKGROUND: Spinacia oleracea is an important dietary vegetable in India and throughout the world and has many beneficial effects. It is cultivated globally. However, its effect on osteoarthritis that mainly targets the cartilage cells remains unknown. In this study we aimed to evaluate the anti-osteoarthritic and chondro-protective effects of SOE on chemically induced osteoarthritis (OA). METHODS: OA was induced by intra-patellar injection of monosodium iodoacetate (MIA) at the knee joint in rats. SOE was then given orally at 250 and 500 mg.kg- 1 day- 1 doses for 28 days to these rats. Anti-osteoarthritic potential of SOE was evaluated by micro-CT, mRNA and protein expression of pro-inflammatory and chondrogenic genes, clinically relevant biomarker's and behavioural experiments. RESULTS: In vitro cell free and cell based assays indicated that SOE acts as a strong anti-oxidant and an anti-inflammatory agent. Histological analysis of knee joints at the end of the experiment by safranin-o and toluidine blue staining established its protective effect. Radiological data corroborated the findings with improvement in the joint space and irregularity of the articular and atrophied femoral condyles and tibial plateau. Micro-CT analysis of sub-chondral bone indicated that SOE had the ability to mitigate OA effects by increasing bone volume to tissue volume (BV/TV) which resulted in decrease of trabecular pattern factor (Tb.Pf) by more than 200%. SOE stimulated chondrogenic marker gene expression with reduction in pro-inflammatory markers. Purified compounds isolated from SOE exhibited increased Sox-9 and Col-II protein expression in articular chondrocytes. Serum and urine analysis indicated that SOE had the potential to down-regulate glutathione S-transferase (GST) activity, clinical markers of osteoarthritis like cartilage oligometric matrix protein (COMP) and CTX-II. Overall, this led to a significant improvement in locomotion and balancing activity in rats as assessed by Open-field and Rota rod test. CONCLUSION: On the basis of in vitro and in vivo experiments performed with Spinacea oleracea extract we can deduce that SOE has the ability to alleviate the MIA induced deleterious effects.