INTRODUCTION:

Osteoporosis (OP) is defined as a systemic skeletal disorder characterized by low bone mass and micro-architectural deterioration of bone tissue with a consequent increase in bone fragility and susceptibility to fracture risk^1-3. It is a serious global health problem associated with high morbidity and mortality, as well as drastic economic burden^4-6. Several drugs with different mechanisms of action have been used for OP with the aim of preventing fracture through inhibition of bone resorption or stimulation of bone formation^7,8. Most currently available approved therapies for OP work by inhibiting the normal breakdown and resorption of bone, such antiresorptive therapies include calcitonin, estrogen replacement therapy, selective estrogen receptor modulators (Raloxifene), bisphosphonates (alendronate sodium and risedronate sodium), and there are new drugs worked by stimulating bone formation such as teriparatide which causes high incidence of hypercalcemia⁹.

Bisphosphonates inhibit farensyl pyrophosphate synthetase, which is responsible for the transformation of geranyl pyrophosphate into farensyl pyrophosphate, and then to geranylgeranyl pyrophosphate which is responsible for osteoclast activation and subsequent bone resorption, through formation of its ruffled borders needed to seal off the bone surface for proper release of proteolytic enzymes and acid that dissolves the underlying bone. However, the safety of oral bisphosphonates has been questioned due to adverse events from the upper gastrointestinal tract, acute phase response, hypocalcaemia, secondary hyperparathyroidism, musculoskeletal pain and osteonecrosis of the jaw^10,11.

Statins are structural analogs of 3-hydroxy-3-methyl glutaryl-coenzyme A and competitively inhibit the HMG-CoA reductase enzyme, which is responsible for the first committed step in sterol biosynthesis. Statins that are approved for use include lovastatin, pravastatin, simvastatin, fluvastatin, cerivastatin, atorvastatin and rosuvastatin. They effectively lower the cholesterol levels, thereby reducing the risk of ischemic heart disease, and stroke. They can also modify endothelial function, control inflammatory response, promote plaque stability, inhibit thrombus formation, reduce platelet aggregation, and maintain a balance between pro thrombotic and fibrinolytic mechanisms^13-18.

They are used in the treatment of hyperlipidemia and act on the same pathway of bisphosphonates but on an earlier step. With respect to the ability of statins to inhibit the HMG-CoA reductase, a key enzyme in the pathways of cholesterol synthesis as well as in the process of activation of osteoclasts, many studies had suggested that statins may have additional important therapeutic effect, these effect known as pleiotropic or cholesterol-independent effects^12,13.

Several in vitro and in vivo animal studies had proven that statins have dual effecton bone formation, as well as antiresorptive effects^13-19.

The aim of this study was to investigate the effect of simvastatin (lipophilic statins) on the bone resorption markers in rats (experimental study).

MATERIALS AND METHODS:

Experimental study animals:

Animals were housed, and were allowed free access to standard dry diet and water. All procedures were in accordance with the National Institute of Health guidelines for the care and use of laboratory animals.

Eighteen adult male Wistar rats were included in the current study. Their weights ranged from 175 to 185 grams, and they were caged in fully ventilated room and exposed to natural daily 12:12 h light–dark cycle. The animals were grouped in poly acrylic cages and maintained under standard laboratory conditions (temperature 25°C±2°C and 50%±5% relative humidity). They were acclimated for 1 week before randomly allocated to groups.

Experimental protocol:

The animals were classified into three groups: group I: included six rats served as control group, group II: included six rats received 7.5mg/kg of dexamethasone intra muscular, weakly for 2 months served as a model group ,group III included six rats received 7.5mg/kg of dexamethasone intra muscular weakly for 2months and received simvastatin daily after 1 month of dexamethasone administration for 4 weeks.

Simvastatin was used at a dose of 8mg/kg/day by oral gavage for 4 weeks²⁰. immediately After 4 weeks,The rats were anesthetized by diethyl ether inhalation before blood obtain, cardiac blood samples were collected from the heart for the three group, and then were centrifuged to separate serum. Serum samples were analyzed for measurement of bone resorptions marker: acid phosphatase (ACP).

Statistical analysis:

The result were analyzed by SPSS 20 statistical package, Excel computer program was used to tabulate the results and represent it graphically. For the quantitative variables which are normally distributed, a paired t-test was used to declare the significant difference at P<0.05. The significant difference between groups was shown using one-way analysis of variance (ANOVA) test at P<0.05.

RESULTS:

The mean serum ACP in group II of rats that received dexamethasone alone for 8 weeks (model group) was significantly elevated to 8.91±1.04µg/L compared to the control group 7.12±0.74µg/L. simvastatin showed a significant reduction of serum ACP to (7.38 ±0.39µg/L) compared to osteoporotic rats group, (8.91±1.04µg/L). The result are shown in Table 1.

Table 1: The mean serum level of ACP

Rat Acp	Normal	Dexamethasone	Simvastatin+ Dexamethasone
1	7	6.9	7.4
2	8.1	9.1	7.1
3	6.3	9.5	7.9
4	7.1	9.2	7.9
5	****	8.9	7.1
6	****	9.9	6.9
Mean ±SD	7.12± 0.74	8.91± 1.04	7.38±0.39

DISCUSSION:

The pleiotropic effect of statins has led clinicians to investigate their potential use among other entities, such as bone metabolism²¹.This perception has mainly been developed by experimental studies, there being a lack of observational studies to clarify the field. The majority of the literature showed an increase in BMD during using of statins²². In fact, the doses used in experimental models which provided a favorable effect were much higher than the doses used in clinical practice²².

The experimental study findings showed that simvastatin reduced the bone resorption markers, denoting that it has antiresorptive effect, on bone. This study revealed increased serum levels of ACP in rats treated with dexamethasone which contributes to high bone turnover rate, being characterized by an increase in bone resorption, leading to bone loss.

The present work showed an increased level of serum TRAP in the model group. Some researchers reported that acid phosphatase is present in bone, spleen, prostate, erythrocytes and platelets. In serum, only bone and erythrocyte isoenzymes of acid phosphatase are insensitive to tartrate, Therefore in the absence of hemolysis, the activity of this isoenzyme, TRAP, can be used as an index of osteoclastic activity²³.

These results coincided with those of El-nabarawi et al who showed that ovariectomy induced high serum TRAP levels as a result of compensation of increased bone turnover²⁴. Furthermore, Sobhani et al showed that changes in serum ACP concentrations represent an increases in bone resorption in the lumbar vertebrae²³.

The relationship between the use of statins and improvement of bone quality reported in the literature is still controversial. Some animal studies have reported positive effects of statins on bone tissue, such as increasing of bone formation²⁵, bone defect healing when applied to the site of injury²⁶, and increasing bone density²⁷. Our results showed that administration of simvastatin to rats with GCs induced osteoporosis produces amelioration in the level of bone turnovers markers.

Our results are compatible with those reported by Lin et al who found that the pleiotropic effect of statins derived its protective effect on the bone by the dual mechanism: suppressing osteoclasts and promoting osteoblastic activities²⁸.

Also, Maeda et. al showed that statins, such as simvastatin and cerivastatin, regulate osteoblast function by increasing the expression of OCN and type I collagen, and by suppressing gene expression of collagenases, such as matrix metallopeptidase (MMP-1) and MMP-13²⁹.

Lipophilic statins, simvastatin, atorvastatin and cerivastatin but not hydrophilic statin, markedly enhance the expression of vascular endothelial growth factor, a bone anabolic factor in osteoblasts²⁴. Uzzan et al ,found in meta-analysis, that statins have a statistically significant positive effect on BMD²¹. In addition, Hughes et al found that the statins of hydrophobic and hydrophilic nature have inhibited osteoclastic action in vitro³⁰, while some other studies have shown that lipophilic agents like simvastatin had better actions^13,19.

The mechanisms of bone anabolism regulated by statins have not been fully elucidated. Some pathways have been identified in which statins may influence bone anabolism (1). Statins increase the levels of bone morphogenetic protein-2 (BMP-2) through the Ras-PI3K-Akt/MAPK signaling pathway, and BMP-2 induces osteoblast differentiation through the Runt-related transcription factor 2 (Runx2) (2). Statins inhibit the mevalonate pathway stopping the synthesis of downstream products, consequently, the FPP and GGPP synthesis are blocked (3). Statins inhibit osteoblast apoptosis via the TGFβ/Smad3 pathway (4). Statins suppress osteoclastogenesis by the OPG/ RANKL/RANK pathway³¹.

Therefore in the future, statins might gain a position among drugs used for the prevention and management of OP. Their anabolic and antiresorptive effects on bone make them an ideal candidate for OP treatment. However, we need further clinical studies with a large number of patients and with a longe period of follow-up.

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Received on 06.03.2021 Modified on 09.05.2021

Research J. Pharm.and Tech 2022; 15(2):525-528.

DOI: 10.52711/0974-360X.2022.00084