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Calcium Phosphate Carrying Simvastatin Enhances Bone Regeneration: A Systematic Review

Abstract

Several studies have aimed to develop alternative therapeutic biomaterials for bone repair. The purpose of this systematic review was to evaluate how statins carried by calcium phosphate affect the formation and regeneration of bone tissue in animal models when compared to other biomaterials or spontaneous healing. This systematic review followed the recommendations of the Cochrane Handbook for Systematic Reviews of Interventions, the PRISMA guidelines, and the Preclinical Systematic Review & Meta-analysis Facility (SyRF). The protocol of this systematic review was registered in PROSPERO (CRD42018091112) and in CAMARADES. In addition, ARRIVE checklists were followed in order to increase the quality and transparency of the search. An electronic search was performed using the MEDLINE/PubMed, Scopus, SciELO, and PROSPERO library databases. The authors used a specific search strategy for each database, and they also conducted a search in the grey literature and cross-references. The eligibility criteria were animal studies, which evaluated bone repair treated with calcium phosphate as a simvastatin carrier. The selection process yielded 8 studies from the 657 retrieved. All manuscripts concluded that locally applied simvastatin carried by calcium phosphate is biocompatible, enhanced bone repair and induced statistically greater bone formation than cloth or calcium phosphate alone. In conclusion, the pertinent pre-clinical studies evidenced the calcium phosphate biocompatibility and its effectiveness in delivering SIM to improve the repair of bone defects. So, clinical trials are encouraged to investigate the impact of SIM associated with calcium phosphate bone graft in repairing bone defect in humans.

Key Words:
statins; simvastatin; calcium phosphate; bone repair; bone regeneration

Resumo

Muitos estudos objetivaram desenvolver biomateriais terapêuticos alternativos para o reparo ósseo. O objetivo desta revisão sistemática foi avaliar o efeito da estatina carreada por fosfato de cálcio na formação e regeneração de tecido ósseo em modelos animais quando comparado com outros biomateriais ou coágulo. Esta revisão sistemática seguiu as recomendações do Cochrane Handbook for Systematic Reviews of Interventions, PRISMA guidelines, e o Preclinical Systematic Review & Meta-analysis Facility (SyRF). O protocolo desta revisão sistemática foi registrado no PROSPERO (CRD42018091112) e no CAMARADES. Além disso, o guia ARRIVE foi utilizado com o objetivo de aumentar a qualidade e transparência do estudo. Uma pesquisa eletrônica foi realizada através do MEDLINE/PubMed, Scopus, SciELO, e biblioteca do PROSPERO. Os autores utilizaram uma estratégia de busca específica para cada base de dados, e uma busca foi conduzida na literatura cinza e nas referências dos artigos selecionados. Os critérios de elegibilidade foram estudos em animais, os quais avaliaram o repara do ósseo tratado com fosfato de cálcio como carreador de estatina. O processo de seleção obteve 8 estudos dos 657 encontrados. Todos os estudos concluíram que a aplicação local da sinvastatina carreada pelo fosfato de cálcio é biocompatível, melhora o reparo ósseo e induz uma formação óssea significantemente maior que coágulo ou fosfato de cálcio sozinho. Em conclusão, os estudos pré-clínicos pertinentes evidenciaram a biocompatibilidade do fosfato de cálcio e sua eficácia na entrega do SIM para melhorar o reparo de defeitos ósseos. Assim, estudos clínicos são encorajados a investigar o impacto do SIM associado ao enxerto ósseo de fosfato de cálcio na reparação de defeito ósseo em humanos.

Introduction

Congenital diseases, infections, traumas and neoplasia can cause bone defect, which is a major problem in medicine and dentistry. Autologous bone grafts is the gold standard for bone regeneration in terms of osteogenesis, osteoconduction, and osteoinduction 11 . Calasans-Maia M, Calasans-Maia J, Santos S, Mavropoulos E, Farina M, Lima I, et al. Short-term in vivo evaluation of zinc-containing calcium phosphate using a normalized procedure. Mater Sci Eng C Mater Biol Appl 2014;41:309-319.. However, these grafts have disadvantages, such as donor-site morbidity, risk of infection, hemorrhage, and chronic pain 22 . Dimitriou R, Mataliotakis GI, Angoules AG, Kanakaris NK, Giannoudis P V. Complications following autologous bone graft harvesting from the iliac crest and using the RIA: A systematic review. Injury 2011;42(SUPPL. 2):S3-15.. For that reason, several studies have aimed to develop alternative therapeutic biomaterials for bone repair 11 . Calasans-Maia M, Calasans-Maia J, Santos S, Mavropoulos E, Farina M, Lima I, et al. Short-term in vivo evaluation of zinc-containing calcium phosphate using a normalized procedure. Mater Sci Eng C Mater Biol Appl 2014;41:309-319.,33 . Suruagy AAP da S, Alves ATNN, Sartoretto SC, Calasans-Maia JA de A, Granjeiro JM, Calasans-Maia MD. Physico-chemical and histomorphometric evaluation of zinc-containing hydroxyapatite in rabbits calvaria. Braz Dent J2016;27:717-726.,44 . Yu W, Sun T-W, Qi C, Ding Z, Zhao H, Zhao S, et al. Evaluation of zinc-doped mesoporous hydroxyapatite microspheres for the construction of a novel biomimetic scaffold optimized for bone augmentation. Int J Nanomedicine 2017;12:2293-2306..

Tissue engineering is based on the following three pillars: biomaterial (scaffold), cells, and bioactive molecules 55 . Lee EJ, Kasper FK, Mikos AG. Biomaterials for tissue engineering. Ann Biomed Eng 2014;42:323-337.. However, because of challenges related to the regulation and translation of cell therapy into clinical practice 66 . A. Ridgway, F. Agbanyo, J. Wang, M. Rosu-Myles, Regulatory Oversight of Cell and Gene Therapy Products in Canada., Adv. Exp. Med. Biol 2015;871:49-71.,77 . Halme DG, Kessler DA. FDA Regulation of Stem-Cell-Based Therapies. N Engl J Med 2006;355:1730-1735.including minimal manipulation and homologous use, may be subjected to a standards-based approach under the Safety of Human Cells, Tissues and Organs for Transplantation Regulations. The manufacture and clinical testing of cell and gene therapy products (CGTPs, there is growing interest in acellular therapy based on drug delivery biomaterials. Consequently, calcium phosphate has been studied as a biomaterial capable of carrying molecules such as bone morphogenetic proteins (BMPs) 88 . Carreira AC, Lojudice FH, Halcsik E, Navarro RD, Sogayar MC, Granjeiro JM. Bone morphogenetic proteins : facts, challenges, and future perspectives. J Dent Res2014;93:335-345., platelet-rich plasma 99 . Malhotra A, Pelletier M, Oliver R, Christou C, Walsh WR. Platelet-Rich Plasma and Bone Defect Healing. Tissue Eng Part A. 2014:2614-2633., strontium ranelate 1010. Querido W, Rossi AL, Farina M. The effects of strontium on bone mineral: A review on current knowledge and microanalytical approaches. Micron2016;80:122-134. and statins 1111. Santana WM de, Sousa DN de, Ferreira VM, Duarte WR. Simvastatin and biphasic calcium phosphate affects bone formation in critical-sized rat calvarial defects. Acta Cir Bras. 2016;31:300-307.. These molecules can stimulate endogenous cells to promote the production of functional bone, resulting in a relatively low-cost product for tissue regeneration therapy 1212. Shah SR, Werlang CA, Kasper FK, Mikos AG. Novel applications of statins for bone regeneration. Natl Sci Rev 2015;2:85-99..

Statins are 3-hydroxy-3-methylglutaryl-coenzyme A (HMG-CoA) reductase inhibitors that are widely used in treating patients with hypercholesterolemia 1313. Endo A. A historical perspective on the discovery of statins. Proc Japan Acad Ser B 2010;86:484-493.. They have also been studied because of their anabolic effects on bone tissue 1414. Papadimitriou K, Karkavelas G, Vouros I, Kessopoulou E, Konstantinidis A. Effects of local application of simvastatin on bone regeneration in femoral bone defects in rabbit. J Cranio-Maxillofacial Surg 2015;43:232-237.,1515. Ayukawa Y, Okamura A, Koyano K. Simvastatin promotes osteogenesis around titanium implants. Clin Oral Implants Res 2004;15:346-350.,1616. Ayukawa Y, Yasukawa E, Moriyama Y, Ogino Y, Wada H, Atsuta I, et al. Local application of statin promotes bone repair through the suppression of osteoclasts and the enhancement of osteoblasts at bone-healing sites in rats. Oral Surgery, Oral Med Oral Pathol Oral Radiol Endodontology 2009;107:336-342.,1717. Mundy G. Stimulation of bone formation in vitro and in rodents by statins. Science1999;286:1946-1949.,1818. Moraschini V, Almeida DCF, Calasans-maia MD. The ability of topical and systemic statins to increase osteogenesis around dental implants : a systematic review of histomorphometric outcomes in animal studies. Int J Oral Maxillofac Surg 2018;47:1070-1078.. Statins increase the expression of BMP-2 1616. Ayukawa Y, Yasukawa E, Moriyama Y, Ogino Y, Wada H, Atsuta I, et al. Local application of statin promotes bone repair through the suppression of osteoclasts and the enhancement of osteoblasts at bone-healing sites in rats. Oral Surgery, Oral Med Oral Pathol Oral Radiol Endodontology 2009;107:336-342.,1717. Mundy G. Stimulation of bone formation in vitro and in rodents by statins. Science1999;286:1946-1949.,1919. Maeda T, Matsunuma A, Kurahashi I, Yanagawa T, Yoshida H, Horiuchi N. Induction of osteoblast differentiation indices by statins in MC3T3-E1 cells. J Cell Biochem 2004;92:458-471. and vascular endothelial growth factor (VEGF) 1919. Maeda T, Matsunuma A, Kurahashi I, Yanagawa T, Yoshida H, Horiuchi N. Induction of osteoblast differentiation indices by statins in MC3T3-E1 cells. J Cell Biochem 2004;92:458-471.. The growth factors BMP-2 and VEGF are of important scientific interest in bone regeneration therapy, but they are expensive and have a short half-life. Therefore, a molecule promoting the endogenous production of BMP-2 and VEGF would be valuable for the field of tissue engineering1212. Shah SR, Werlang CA, Kasper FK, Mikos AG. Novel applications of statins for bone regeneration. Natl Sci Rev 2015;2:85-99..

Statins can be administered systemically or locally within a vehicle or carrier. Studies using systemic administration indicate that severe muscular side effects are rare; however, mild side effects, such as myopathic symptoms, are common 2020. Sinzinger H, Wolfram R, Peskar BA. Muscular side effects of statins. J Cardiovasc Pharmacol2002;40:163-171.. For this reason, topical administration is recommended for bone regeneration, since local application allows a controlled, gradual release of the molecule, increasing its bioavailability and effectiveness 1212. Shah SR, Werlang CA, Kasper FK, Mikos AG. Novel applications of statins for bone regeneration. Natl Sci Rev 2015;2:85-99.,1414. Papadimitriou K, Karkavelas G, Vouros I, Kessopoulou E, Konstantinidis A. Effects of local application of simvastatin on bone regeneration in femoral bone defects in rabbit. J Cranio-Maxillofacial Surg 2015;43:232-237.. Therefore, for local delivery strategy calcium phosphate can be used as a carrier. In addition, this material is biocompatible, osteoconductive, act as a space maintainer for bone formation and exhibit varying degrees of resorbability 33 . Suruagy AAP da S, Alves ATNN, Sartoretto SC, Calasans-Maia JA de A, Granjeiro JM, Calasans-Maia MD. Physico-chemical and histomorphometric evaluation of zinc-containing hydroxyapatite in rabbits calvaria. Braz Dent J2016;27:717-726.,2121. Shepherd JH, Shepherd D V, Best SM. Substituted hydroxyapatites for bone repair. J Mater Sci Mater Med2012;23:2335-2347.. For that reason, the aim of this systematic review (SR) is to evaluate the influence of calcium phosphate as a statin carrier on bone repair in animal studies, to provide information on potential clinical application. The following question was stated: How do statins carried by calcium phosphate affect the formation and regeneration of bone tissue in animal studies?

Material and Methods

The protocol of this systematic review (SR) was recorded in PROSPERO database under number CRD42018091112 in CAMARADES at http://syrf.org.uk/protocols/. This SR followed the recommendations of the Cochrane Handbook for Systematic Reviews of Interventions [22]. In order to increase the quality and transparency of the study, the PRISMA [23] and ARRIVE checklists 2222. Kilkenny C, Browne WJ, Cuthill IC, Emerson M, Altman DG. Improving bioscience research reporting: The arrive guidelines for reporting animal research. PLoS Biol2010;8. were followed.

Focused Question (Based On PICO Criteria)

How do statins carried by calcium phosphate (I) affect the formation and regeneration of bone tissue (O) in animal models (P) when compared to other biomaterials or spontaneous healing (C)?

Outcome Measures

The primary outcome measures were bone formation through histological and histomorphometrical analysis. The secondary outcome variables were the presence of biomaterial and biocompatibility.

Search Strategy

An electronic search was carried out in the MEDLINE/PubMed, Scopus, SciELO, and PROSPERO library databases up to October 2018. There was no date restriction, but only studies in English, Portuguese, Spanish, or French were included. In addition, a manual electronic search of some periodical journals was performed. Unpublished studies (i.e., grey literature) were searched for in the Grey Literature Report and OpenGrey databases. Searches in the references of the included studies (i.e., cross-referencing) were also conducted. A specific search strategy was used for each database, according to its characteristics (Box 1). Aiming for in vivo studies, a search filter was developed 2323. Hooijmans CR, Tillema A, Leenaars M, Ritskes-hoitinga M. Original Article Enhancing search efficiency by means of a search filter for finding all studies on animal experimentation in PubMed. Lab animails 2010;44:170-175. that could detect 7% more records than the regular method, the PubMed Limit: Animals.

Box 1
Search strategy

Eligibility Criteria

Inclusion Criteria

Pre-clinical and experimental studies that evaluated bone repair through calcium phosphate biomaterial as a carrier for statins compared to other biomaterials or blood clots

Exclusion Criteria

The exclusion criteria were manuscripts involving in vitro studies, syntheses, characterization of biomaterials, and reviews; studies in animals with systemic disorders or infections, such as osteoporosis or osteopenia; studies involving biomaterial associated/coating implant and alloy surface, systemic administration of statins, biomaterial associated with substances other than statins, cells or growth factors, statin ectopic injection; or studies having an outcome unrelated to bone formation, osteoconduction, bioresorption, or biocompatibility.

Study Selection, Screening Process and Data Extraction

The process of searching and selecting the studies was conducted in duplicate by two authors (M.D.C.M. and R.C). First, titles and abstracts were carefully evaluated. After the first evaluation, potential articles were carefully assessed according to the eligibility criteria of this review. Possible disagreements were resolved through a consensus between the authors. When necessary, the authors of the included studies were contacted by email for clarification.

The following data were extracted from the papers by two reviewers (G.P. and R.C.): DOI, authors, year, animal model, sample size, experimental period, biomaterials used, location of biomaterial implantation, analysis method, outcome, and main conclusion.

Assessment of Quality and the Risk of Bias

The quality assessment was done by two reviewers (R.C. and M.D.C.M.), using the ARRIVE guideline checklist 2222. Kilkenny C, Browne WJ, Cuthill IC, Emerson M, Altman DG. Improving bioscience research reporting: The arrive guidelines for reporting animal research. PLoS Biol2010;8.. According to these guidelines, a checklist consisting of 26 items was developed; based on a previous study 2424. Stadlinger B, Pourmand P, Locher MC, Schulz MC. Systematic review of animal models for the study of implant integration, assessing the influence of material, surface and design. J Clin Periodontol2012;39(SUPPL.12):28-36., each criterion was graded as “0” (not reported or not performed) or “1” (reported). A final score was then recorded for each study.

The risk of bias (ROB) was performed by two reviewing authors (M.D.C.M. and R.C.) using the risk-of-bias tool developed by the Systematic Review Center for Laboratory Animal Experimentation (SYRCLE) 2525. Hooijmans CR, Rovers MM, de Vries RB, Leenaars M, Ritskes-Hoitinga M. SYRCLE’s risk of bias tool for animal studies. BMC Med Res Methodol2014;14:1-9.. The goal of this tool is to assess the quality of the applied methodology by classifying the papers as having a low, high, or unclear risk of bias.

Statistical Analysis

Means and standard deviations were extracted from the selected studies, and descriptive analyses were conducted. The software Microsoft Office Excel (2010) was used for descriptive statistics. A pairwise meta-analysis could not be conducted because of the heterogeneity between studies, which used different animal models, carrier biomaterials, administration sites, doses, and follow-up durations.

Results

Literature Search

The initial search resulted in 657 articles, comprising 371 titles from MEDLINE/PubMed, 12 from SciELO/Bireme, 270 from Scopus, three from PROSPERO, and one from cross-referencing. The search in the grey literature did not yield the inclusion of any additional studies. Duplicates were removed, and after screening the titles and abstracts, 14 studies were selected. Following the screening of the full text of the 14 selected studies, 6 manuscripts were excluded because they did not meet the eligibility criteria. As a result, 8 studies 1111. Santana WM de, Sousa DN de, Ferreira VM, Duarte WR. Simvastatin and biphasic calcium phosphate affects bone formation in critical-sized rat calvarial defects. Acta Cir Bras. 2016;31:300-307.,2626. Adah F, Benghuzzi H, Tucci M, Russell G, England B. Effects of sustained release of statin by means of tricalcium phosphate lysine delivery system in defect and segmental femoral injuries on certain biochemical markers in vivo. Biomed Sci Instrum . 2006;42:126-35.,2727. Gao S, Shiota M, Fujii M, Chen K, Shimogishi M. Combination of simvastatin and hydroxyapatite fiber induces bone augmentation. Open J Regen Med 2013;2:53-60.,2828. Nyan M, Miyahara T, Noritake K, Hao J, Rodriguez R, Kuroda S, et al. Molecular and tissue responses in the healing of rat calvarial defects after local application of simvastatin combined with alpha tricalcium phosphate. J Biomed Mater Res Part B Appl Biomater 2010;93:65-73.,2929. Nyan M, Sato D, Kihara H, Machida T, Ohya K, Kasugai S. Effects of the combination with alpha-tricalcium phosphate and simvastatin on bone regeneration. Clin Oral Implants Res 2009;20:280-287.,3030. Rojbani H, Nyan M, Ohya K, Kasugai S. Evaluation of the osteoconductivity of α-tricalcium phosphate, β-tricalcium phosphate, and hydroxyapatite combined with or without simvastatin in rat calvarial defect. J Biomed Mater Res A 2011;98:488-498.,3131. Shahrezaie M, Moshiri A, Shekarchi B, Oryan A, Maffulli N, Parvizi J. Effectiveness of tissue engineered three-dimensional bioactive graft on bone healing and regeneration: an in vivo study with significant clinical value. J Tissue Eng Regen Med2017;1-15.,3232. Yin H, Li Y-G, Si M, Li J-M. Simvastatin-loaded macroporous calcium phosphate cement: preparation, in vitro characterization, and evaluation of in vivo performance. J Biomed Mater Res A 2012;100:2991-3000. were included in this systematic review (Fig 1).

Fig. 1
Flow diagram of literature selection process. * E.g. reviews, letters and comments.

Assessment of Quality and the Risk of Bias

With regard to quality assessment, the mean and median score of the studies were 11.75 (±2.05) and 12.5, respectively. The highest score was 15 points 1111. Santana WM de, Sousa DN de, Ferreira VM, Duarte WR. Simvastatin and biphasic calcium phosphate affects bone formation in critical-sized rat calvarial defects. Acta Cir Bras. 2016;31:300-307., and the lowest was 9 2828. Nyan M, Miyahara T, Noritake K, Hao J, Rodriguez R, Kuroda S, et al. Molecular and tissue responses in the healing of rat calvarial defects after local application of simvastatin combined with alpha tricalcium phosphate. J Biomed Mater Res Part B Appl Biomater 2010;93:65-73.,3030. Rojbani H, Nyan M, Ohya K, Kasugai S. Evaluation of the osteoconductivity of α-tricalcium phosphate, β-tricalcium phosphate, and hydroxyapatite combined with or without simvastatin in rat calvarial defect. J Biomed Mater Res A 2011;98:488-498.. None of the studies presented sample size calculations or baseline animal health data, and none mentioned blinding of accessor. In addition, none of the manuscripts described the implications of experimental methods for replacement, refinement, or reduction (the 3Rs) or discussed the relevance of the study to human biology. Regarding statistical methods, one study was qualitative, so it did not present statistics 2626. Adah F, Benghuzzi H, Tucci M, Russell G, England B. Effects of sustained release of statin by means of tricalcium phosphate lysine delivery system in defect and segmental femoral injuries on certain biochemical markers in vivo. Biomed Sci Instrum . 2006;42:126-35.. All of the studies clearly defined the outcomes assessed and the study design. However, only one study mentioned parameters such as biomaterial implant randomization 2727. Gao S, Shiota M, Fujii M, Chen K, Shimogishi M. Combination of simvastatin and hydroxyapatite fiber induces bone augmentation. Open J Regen Med 2013;2:53-60. and house and husbandry 1111. Santana WM de, Sousa DN de, Ferreira VM, Duarte WR. Simvastatin and biphasic calcium phosphate affects bone formation in critical-sized rat calvarial defects. Acta Cir Bras. 2016;31:300-307. (Table 1).

Table 1
Quality assessment - Arrive-based score

With respect to ROB, in 75% of the manuscripts all animals were included in the analysis. The baseline condition was reported to be the same for every group in half of the studies, considering that there was no disease induction. Seven out of eight studies reported all of the expected outcomes. None of the manuscripts were described as low risk for blinding in detection bias or sequence generation in selection bias. However, one study claimed that the allocation to different groups was done randomly 2727. Gao S, Shiota M, Fujii M, Chen K, Shimogishi M. Combination of simvastatin and hydroxyapatite fiber induces bone augmentation. Open J Regen Med 2013;2:53-60..

Study Characteristics

In the eligible manuscripts, the experimental animal models used were rabbits2727. Gao S, Shiota M, Fujii M, Chen K, Shimogishi M. Combination of simvastatin and hydroxyapatite fiber induces bone augmentation. Open J Regen Med 2013;2:53-60.,3131. Shahrezaie M, Moshiri A, Shekarchi B, Oryan A, Maffulli N, Parvizi J. Effectiveness of tissue engineered three-dimensional bioactive graft on bone healing and regeneration: an in vivo study with significant clinical value. J Tissue Eng Regen Med2017;1-15.,3232. Yin H, Li Y-G, Si M, Li J-M. Simvastatin-loaded macroporous calcium phosphate cement: preparation, in vitro characterization, and evaluation of in vivo performance. J Biomed Mater Res A 2012;100:2991-3000. and rats 1111. Santana WM de, Sousa DN de, Ferreira VM, Duarte WR. Simvastatin and biphasic calcium phosphate affects bone formation in critical-sized rat calvarial defects. Acta Cir Bras. 2016;31:300-307.,2626. Adah F, Benghuzzi H, Tucci M, Russell G, England B. Effects of sustained release of statin by means of tricalcium phosphate lysine delivery system in defect and segmental femoral injuries on certain biochemical markers in vivo. Biomed Sci Instrum . 2006;42:126-35.,2828. Nyan M, Miyahara T, Noritake K, Hao J, Rodriguez R, Kuroda S, et al. Molecular and tissue responses in the healing of rat calvarial defects after local application of simvastatin combined with alpha tricalcium phosphate. J Biomed Mater Res Part B Appl Biomater 2010;93:65-73.,2929. Nyan M, Sato D, Kihara H, Machida T, Ohya K, Kasugai S. Effects of the combination with alpha-tricalcium phosphate and simvastatin on bone regeneration. Clin Oral Implants Res 2009;20:280-287.,3030. Rojbani H, Nyan M, Ohya K, Kasugai S. Evaluation of the osteoconductivity of α-tricalcium phosphate, β-tricalcium phosphate, and hydroxyapatite combined with or without simvastatin in rat calvarial defect. J Biomed Mater Res A 2011;98:488-498., with a total of 291 animals (72 rabbits and 219 rats). The sample size varied from 3 to 9 (Fig 2). The experimental period was from 3 to 56 days. The calcium phosphate used in the eligible manuscripts was tricalcium phosphate lysine 2626. Adah F, Benghuzzi H, Tucci M, Russell G, England B. Effects of sustained release of statin by means of tricalcium phosphate lysine delivery system in defect and segmental femoral injuries on certain biochemical markers in vivo. Biomed Sci Instrum . 2006;42:126-35.; alpha-tricalcium phosphate (α-TCP)2828. Nyan M, Miyahara T, Noritake K, Hao J, Rodriguez R, Kuroda S, et al. Molecular and tissue responses in the healing of rat calvarial defects after local application of simvastatin combined with alpha tricalcium phosphate. J Biomed Mater Res Part B Appl Biomater 2010;93:65-73.,2929. Nyan M, Sato D, Kihara H, Machida T, Ohya K, Kasugai S. Effects of the combination with alpha-tricalcium phosphate and simvastatin on bone regeneration. Clin Oral Implants Res 2009;20:280-287.; α-TCP, beta-tricalcium phosphate (β-TCP) and hydroxyapatite (HA) 3030. Rojbani H, Nyan M, Ohya K, Kasugai S. Evaluation of the osteoconductivity of α-tricalcium phosphate, β-tricalcium phosphate, and hydroxyapatite combined with or without simvastatin in rat calvarial defect. J Biomed Mater Res A 2011;98:488-498.; macroporous calcium phosphate bone cements 3232. Yin H, Li Y-G, Si M, Li J-M. Simvastatin-loaded macroporous calcium phosphate cement: preparation, in vitro characterization, and evaluation of in vivo performance. J Biomed Mater Res A 2012;100:2991-3000.; hydroxyapatite fiber (HAF) 2727. Gao S, Shiota M, Fujii M, Chen K, Shimogishi M. Combination of simvastatin and hydroxyapatite fiber induces bone augmentation. Open J Regen Med 2013;2:53-60.; biphasic calcium phosphate (BCP) 1111. Santana WM de, Sousa DN de, Ferreira VM, Duarte WR. Simvastatin and biphasic calcium phosphate affects bone formation in critical-sized rat calvarial defects. Acta Cir Bras. 2016;31:300-307.; and nanostructured-hydroxyapatite (nHA) 3131. Shahrezaie M, Moshiri A, Shekarchi B, Oryan A, Maffulli N, Parvizi J. Effectiveness of tissue engineered three-dimensional bioactive graft on bone healing and regeneration: an in vivo study with significant clinical value. J Tissue Eng Regen Med2017;1-15.. In this study, the presentation form of the biomaterials used was a cylinder 2626. Adah F, Benghuzzi H, Tucci M, Russell G, England B. Effects of sustained release of statin by means of tricalcium phosphate lysine delivery system in defect and segmental femoral injuries on certain biochemical markers in vivo. Biomed Sci Instrum . 2006;42:126-35.; particles 500-700 µm2828. Nyan M, Miyahara T, Noritake K, Hao J, Rodriguez R, Kuroda S, et al. Molecular and tissue responses in the healing of rat calvarial defects after local application of simvastatin combined with alpha tricalcium phosphate. J Biomed Mater Res Part B Appl Biomater 2010;93:65-73.,2929. Nyan M, Sato D, Kihara H, Machida T, Ohya K, Kasugai S. Effects of the combination with alpha-tricalcium phosphate and simvastatin on bone regeneration. Clin Oral Implants Res 2009;20:280-287.; particles 500-750 µm 3030. Rojbani H, Nyan M, Ohya K, Kasugai S. Evaluation of the osteoconductivity of α-tricalcium phosphate, β-tricalcium phosphate, and hydroxyapatite combined with or without simvastatin in rat calvarial defect. J Biomed Mater Res A 2011;98:488-498.; cement paste and cylinders 3232. Yin H, Li Y-G, Si M, Li J-M. Simvastatin-loaded macroporous calcium phosphate cement: preparation, in vitro characterization, and evaluation of in vivo performance. J Biomed Mater Res A 2012;100:2991-3000.; fibers 2727. Gao S, Shiota M, Fujii M, Chen K, Shimogishi M. Combination of simvastatin and hydroxyapatite fiber induces bone augmentation. Open J Regen Med 2013;2:53-60.; particles 1111. Santana WM de, Sousa DN de, Ferreira VM, Duarte WR. Simvastatin and biphasic calcium phosphate affects bone formation in critical-sized rat calvarial defects. Acta Cir Bras. 2016;31:300-307., and powder 3131. Shahrezaie M, Moshiri A, Shekarchi B, Oryan A, Maffulli N, Parvizi J. Effectiveness of tissue engineered three-dimensional bioactive graft on bone healing and regeneration: an in vivo study with significant clinical value. J Tissue Eng Regen Med2017;1-15.. The biomaterial grafts were inserted into the region of the femur, with perforation around 2 mm 2626. Adah F, Benghuzzi H, Tucci M, Russell G, England B. Effects of sustained release of statin by means of tricalcium phosphate lysine delivery system in defect and segmental femoral injuries on certain biochemical markers in vivo. Biomed Sci Instrum . 2006;42:126-35., 3 mm 3131. Shahrezaie M, Moshiri A, Shekarchi B, Oryan A, Maffulli N, Parvizi J. Effectiveness of tissue engineered three-dimensional bioactive graft on bone healing and regeneration: an in vivo study with significant clinical value. J Tissue Eng Regen Med2017;1-15., and 5 mm 3232. Yin H, Li Y-G, Si M, Li J-M. Simvastatin-loaded macroporous calcium phosphate cement: preparation, in vitro characterization, and evaluation of in vivo performance. J Biomed Mater Res A 2012;100:2991-3000.; the muscle 3232. Yin H, Li Y-G, Si M, Li J-M. Simvastatin-loaded macroporous calcium phosphate cement: preparation, in vitro characterization, and evaluation of in vivo performance. J Biomed Mater Res A 2012;100:2991-3000.; and the bilateral calvaria 5 mm defects 1111. Santana WM de, Sousa DN de, Ferreira VM, Duarte WR. Simvastatin and biphasic calcium phosphate affects bone formation in critical-sized rat calvarial defects. Acta Cir Bras. 2016;31:300-307.,2727. Gao S, Shiota M, Fujii M, Chen K, Shimogishi M. Combination of simvastatin and hydroxyapatite fiber induces bone augmentation. Open J Regen Med 2013;2:53-60.,2828. Nyan M, Miyahara T, Noritake K, Hao J, Rodriguez R, Kuroda S, et al. Molecular and tissue responses in the healing of rat calvarial defects after local application of simvastatin combined with alpha tricalcium phosphate. J Biomed Mater Res Part B Appl Biomater 2010;93:65-73.,2929. Nyan M, Sato D, Kihara H, Machida T, Ohya K, Kasugai S. Effects of the combination with alpha-tricalcium phosphate and simvastatin on bone regeneration. Clin Oral Implants Res 2009;20:280-287.,3030. Rojbani H, Nyan M, Ohya K, Kasugai S. Evaluation of the osteoconductivity of α-tricalcium phosphate, β-tricalcium phosphate, and hydroxyapatite combined with or without simvastatin in rat calvarial defect. J Biomed Mater Res A 2011;98:488-498..

Fig. 2
Study characteristics. n*= sample size median per animal model

Plasma statin concentration was not analyzed in all studies. The method of analysis most often applied was histological analysis 1111. Santana WM de, Sousa DN de, Ferreira VM, Duarte WR. Simvastatin and biphasic calcium phosphate affects bone formation in critical-sized rat calvarial defects. Acta Cir Bras. 2016;31:300-307.,2727. Gao S, Shiota M, Fujii M, Chen K, Shimogishi M. Combination of simvastatin and hydroxyapatite fiber induces bone augmentation. Open J Regen Med 2013;2:53-60.,2828. Nyan M, Miyahara T, Noritake K, Hao J, Rodriguez R, Kuroda S, et al. Molecular and tissue responses in the healing of rat calvarial defects after local application of simvastatin combined with alpha tricalcium phosphate. J Biomed Mater Res Part B Appl Biomater 2010;93:65-73.,2929. Nyan M, Sato D, Kihara H, Machida T, Ohya K, Kasugai S. Effects of the combination with alpha-tricalcium phosphate and simvastatin on bone regeneration. Clin Oral Implants Res 2009;20:280-287.,3030. Rojbani H, Nyan M, Ohya K, Kasugai S. Evaluation of the osteoconductivity of α-tricalcium phosphate, β-tricalcium phosphate, and hydroxyapatite combined with or without simvastatin in rat calvarial defect. J Biomed Mater Res A 2011;98:488-498.,3131. Shahrezaie M, Moshiri A, Shekarchi B, Oryan A, Maffulli N, Parvizi J. Effectiveness of tissue engineered three-dimensional bioactive graft on bone healing and regeneration: an in vivo study with significant clinical value. J Tissue Eng Regen Med2017;1-15.,3232. Yin H, Li Y-G, Si M, Li J-M. Simvastatin-loaded macroporous calcium phosphate cement: preparation, in vitro characterization, and evaluation of in vivo performance. J Biomed Mater Res A 2012;100:2991-3000. and histomorphometrical evaluation 1111. Santana WM de, Sousa DN de, Ferreira VM, Duarte WR. Simvastatin and biphasic calcium phosphate affects bone formation in critical-sized rat calvarial defects. Acta Cir Bras. 2016;31:300-307.,2626. Adah F, Benghuzzi H, Tucci M, Russell G, England B. Effects of sustained release of statin by means of tricalcium phosphate lysine delivery system in defect and segmental femoral injuries on certain biochemical markers in vivo. Biomed Sci Instrum . 2006;42:126-35.,2727. Gao S, Shiota M, Fujii M, Chen K, Shimogishi M. Combination of simvastatin and hydroxyapatite fiber induces bone augmentation. Open J Regen Med 2013;2:53-60.,2828. Nyan M, Miyahara T, Noritake K, Hao J, Rodriguez R, Kuroda S, et al. Molecular and tissue responses in the healing of rat calvarial defects after local application of simvastatin combined with alpha tricalcium phosphate. J Biomed Mater Res Part B Appl Biomater 2010;93:65-73.,2929. Nyan M, Sato D, Kihara H, Machida T, Ohya K, Kasugai S. Effects of the combination with alpha-tricalcium phosphate and simvastatin on bone regeneration. Clin Oral Implants Res 2009;20:280-287.,3030. Rojbani H, Nyan M, Ohya K, Kasugai S. Evaluation of the osteoconductivity of α-tricalcium phosphate, β-tricalcium phosphate, and hydroxyapatite combined with or without simvastatin in rat calvarial defect. J Biomed Mater Res A 2011;98:488-498.,3131. Shahrezaie M, Moshiri A, Shekarchi B, Oryan A, Maffulli N, Parvizi J. Effectiveness of tissue engineered three-dimensional bioactive graft on bone healing and regeneration: an in vivo study with significant clinical value. J Tissue Eng Regen Med2017;1-15.,3232. Yin H, Li Y-G, Si M, Li J-M. Simvastatin-loaded macroporous calcium phosphate cement: preparation, in vitro characterization, and evaluation of in vivo performance. J Biomed Mater Res A 2012;100:2991-3000., followed by micro-CT evaluation 2727. Gao S, Shiota M, Fujii M, Chen K, Shimogishi M. Combination of simvastatin and hydroxyapatite fiber induces bone augmentation. Open J Regen Med 2013;2:53-60.,2929. Nyan M, Sato D, Kihara H, Machida T, Ohya K, Kasugai S. Effects of the combination with alpha-tricalcium phosphate and simvastatin on bone regeneration. Clin Oral Implants Res 2009;20:280-287.,3030. Rojbani H, Nyan M, Ohya K, Kasugai S. Evaluation of the osteoconductivity of α-tricalcium phosphate, β-tricalcium phosphate, and hydroxyapatite combined with or without simvastatin in rat calvarial defect. J Biomed Mater Res A 2011;98:488-498.,3131. Shahrezaie M, Moshiri A, Shekarchi B, Oryan A, Maffulli N, Parvizi J. Effectiveness of tissue engineered three-dimensional bioactive graft on bone healing and regeneration: an in vivo study with significant clinical value. J Tissue Eng Regen Med2017;1-15. and radiological and thermograph analysis 3131. Shahrezaie M, Moshiri A, Shekarchi B, Oryan A, Maffulli N, Parvizi J. Effectiveness of tissue engineered three-dimensional bioactive graft on bone healing and regeneration: an in vivo study with significant clinical value. J Tissue Eng Regen Med2017;1-15., considering overlapping parameters. In addition, the parameters considered were newly formed bone 1111. Santana WM de, Sousa DN de, Ferreira VM, Duarte WR. Simvastatin and biphasic calcium phosphate affects bone formation in critical-sized rat calvarial defects. Acta Cir Bras. 2016;31:300-307.,2626. Adah F, Benghuzzi H, Tucci M, Russell G, England B. Effects of sustained release of statin by means of tricalcium phosphate lysine delivery system in defect and segmental femoral injuries on certain biochemical markers in vivo. Biomed Sci Instrum . 2006;42:126-35.,2727. Gao S, Shiota M, Fujii M, Chen K, Shimogishi M. Combination of simvastatin and hydroxyapatite fiber induces bone augmentation. Open J Regen Med 2013;2:53-60.,2828. Nyan M, Miyahara T, Noritake K, Hao J, Rodriguez R, Kuroda S, et al. Molecular and tissue responses in the healing of rat calvarial defects after local application of simvastatin combined with alpha tricalcium phosphate. J Biomed Mater Res Part B Appl Biomater 2010;93:65-73.,2929. Nyan M, Sato D, Kihara H, Machida T, Ohya K, Kasugai S. Effects of the combination with alpha-tricalcium phosphate and simvastatin on bone regeneration. Clin Oral Implants Res 2009;20:280-287.,3030. Rojbani H, Nyan M, Ohya K, Kasugai S. Evaluation of the osteoconductivity of α-tricalcium phosphate, β-tricalcium phosphate, and hydroxyapatite combined with or without simvastatin in rat calvarial defect. J Biomed Mater Res A 2011;98:488-498.,3131. Shahrezaie M, Moshiri A, Shekarchi B, Oryan A, Maffulli N, Parvizi J. Effectiveness of tissue engineered three-dimensional bioactive graft on bone healing and regeneration: an in vivo study with significant clinical value. J Tissue Eng Regen Med2017;1-15.,3232. Yin H, Li Y-G, Si M, Li J-M. Simvastatin-loaded macroporous calcium phosphate cement: preparation, in vitro characterization, and evaluation of in vivo performance. J Biomed Mater Res A 2012;100:2991-3000.; bioresorption 3030. Rojbani H, Nyan M, Ohya K, Kasugai S. Evaluation of the osteoconductivity of α-tricalcium phosphate, β-tricalcium phosphate, and hydroxyapatite combined with or without simvastatin in rat calvarial defect. J Biomed Mater Res A 2011;98:488-498.,3131. Shahrezaie M, Moshiri A, Shekarchi B, Oryan A, Maffulli N, Parvizi J. Effectiveness of tissue engineered three-dimensional bioactive graft on bone healing and regeneration: an in vivo study with significant clinical value. J Tissue Eng Regen Med2017;1-15.; biocompatibility 2929. Nyan M, Sato D, Kihara H, Machida T, Ohya K, Kasugai S. Effects of the combination with alpha-tricalcium phosphate and simvastatin on bone regeneration. Clin Oral Implants Res 2009;20:280-287.,3131. Shahrezaie M, Moshiri A, Shekarchi B, Oryan A, Maffulli N, Parvizi J. Effectiveness of tissue engineered three-dimensional bioactive graft on bone healing and regeneration: an in vivo study with significant clinical value. J Tissue Eng Regen Med2017;1-15.,3232. Yin H, Li Y-G, Si M, Li J-M. Simvastatin-loaded macroporous calcium phosphate cement: preparation, in vitro characterization, and evaluation of in vivo performance. J Biomed Mater Res A 2012;100:2991-3000.; bone mineral content and bone mineral density 2929. Nyan M, Sato D, Kihara H, Machida T, Ohya K, Kasugai S. Effects of the combination with alpha-tricalcium phosphate and simvastatin on bone regeneration. Clin Oral Implants Res 2009;20:280-287.,3232. Yin H, Li Y-G, Si M, Li J-M. Simvastatin-loaded macroporous calcium phosphate cement: preparation, in vitro characterization, and evaluation of in vivo performance. J Biomed Mater Res A 2012;100:2991-3000.; simvastatin (SIM) release in vitro 2727. Gao S, Shiota M, Fujii M, Chen K, Shimogishi M. Combination of simvastatin and hydroxyapatite fiber induces bone augmentation. Open J Regen Med 2013;2:53-60.,2828. Nyan M, Miyahara T, Noritake K, Hao J, Rodriguez R, Kuroda S, et al. Molecular and tissue responses in the healing of rat calvarial defects after local application of simvastatin combined with alpha tricalcium phosphate. J Biomed Mater Res Part B Appl Biomater 2010;93:65-73.,3232. Yin H, Li Y-G, Si M, Li J-M. Simvastatin-loaded macroporous calcium phosphate cement: preparation, in vitro characterization, and evaluation of in vivo performance. J Biomed Mater Res A 2012;100:2991-3000.; morphology of the vital organs 2626. Adah F, Benghuzzi H, Tucci M, Russell G, England B. Effects of sustained release of statin by means of tricalcium phosphate lysine delivery system in defect and segmental femoral injuries on certain biochemical markers in vivo. Biomed Sci Instrum . 2006;42:126-35.; BMP, TGF, and VEGF gene expression 2828. Nyan M, Miyahara T, Noritake K, Hao J, Rodriguez R, Kuroda S, et al. Molecular and tissue responses in the healing of rat calvarial defects after local application of simvastatin combined with alpha tricalcium phosphate. J Biomed Mater Res Part B Appl Biomater 2010;93:65-73.; and bone labeling with calcein at 7 days and tetracycline at 1 day 3030. Rojbani H, Nyan M, Ohya K, Kasugai S. Evaluation of the osteoconductivity of α-tricalcium phosphate, β-tricalcium phosphate, and hydroxyapatite combined with or without simvastatin in rat calvarial defect. J Biomed Mater Res A 2011;98:488-498..

Biocompatibility

The biocompatibility studies used rabbits as their animal models 3131. Shahrezaie M, Moshiri A, Shekarchi B, Oryan A, Maffulli N, Parvizi J. Effectiveness of tissue engineered three-dimensional bioactive graft on bone healing and regeneration: an in vivo study with significant clinical value. J Tissue Eng Regen Med2017;1-15.,3232. Yin H, Li Y-G, Si M, Li J-M. Simvastatin-loaded macroporous calcium phosphate cement: preparation, in vitro characterization, and evaluation of in vivo performance. J Biomed Mater Res A 2012;100:2991-3000. . Yin et al. used macroporous calcium phosphate cement (CPC), to be compared with CPC associated to simvastatin 3232. Yin H, Li Y-G, Si M, Li J-M. Simvastatin-loaded macroporous calcium phosphate cement: preparation, in vitro characterization, and evaluation of in vivo performance. J Biomed Mater Res A 2012;100:2991-3000., and Shahrezaie et al. used nanostructured hydroxyapatite particles with and without simvastatin 3131. Shahrezaie M, Moshiri A, Shekarchi B, Oryan A, Maffulli N, Parvizi J. Effectiveness of tissue engineered three-dimensional bioactive graft on bone healing and regeneration: an in vivo study with significant clinical value. J Tissue Eng Regen Med2017;1-15.. The reported results indicate that small amounts (1 wt%) of SIM associated with the CPCs did not affect the physico-chemical properties or biocompatibility 3232. Yin H, Li Y-G, Si M, Li J-M. Simvastatin-loaded macroporous calcium phosphate cement: preparation, in vitro characterization, and evaluation of in vivo performance. J Biomed Mater Res A 2012;100:2991-3000., and nHA and SIM particles, when used to fill bone defect, were biocompatible with and beneficial to bone healing 3131. Shahrezaie M, Moshiri A, Shekarchi B, Oryan A, Maffulli N, Parvizi J. Effectiveness of tissue engineered three-dimensional bioactive graft on bone healing and regeneration: an in vivo study with significant clinical value. J Tissue Eng Regen Med2017;1-15.. Manuscripts that didn’t focus on biocompatibility but that evaluated the inflammatory response described no or slight inflammatory reaction with low concentrations of SIM 1111. Santana WM de, Sousa DN de, Ferreira VM, Duarte WR. Simvastatin and biphasic calcium phosphate affects bone formation in critical-sized rat calvarial defects. Acta Cir Bras. 2016;31:300-307.,27,27. Gao S, Shiota M, Fujii M, Chen K, Shimogishi M. Combination of simvastatin and hydroxyapatite fiber induces bone augmentation. Open J Regen Med 2013;2:53-60. 2828. Nyan M, Miyahara T, Noritake K, Hao J, Rodriguez R, Kuroda S, et al. Molecular and tissue responses in the healing of rat calvarial defects after local application of simvastatin combined with alpha tricalcium phosphate. J Biomed Mater Res Part B Appl Biomater 2010;93:65-73.,2929. Nyan M, Sato D, Kihara H, Machida T, Ohya K, Kasugai S. Effects of the combination with alpha-tricalcium phosphate and simvastatin on bone regeneration. Clin Oral Implants Res 2009;20:280-287.,3030. Rojbani H, Nyan M, Ohya K, Kasugai S. Evaluation of the osteoconductivity of α-tricalcium phosphate, β-tricalcium phosphate, and hydroxyapatite combined with or without simvastatin in rat calvarial defect. J Biomed Mater Res A 2011;98:488-498.,3131. Shahrezaie M, Moshiri A, Shekarchi B, Oryan A, Maffulli N, Parvizi J. Effectiveness of tissue engineered three-dimensional bioactive graft on bone healing and regeneration: an in vivo study with significant clinical value. J Tissue Eng Regen Med2017;1-15.,3232. Yin H, Li Y-G, Si M, Li J-M. Simvastatin-loaded macroporous calcium phosphate cement: preparation, in vitro characterization, and evaluation of in vivo performance. J Biomed Mater Res A 2012;100:2991-3000., which decreased over the experimental period in all studies, regardless of the implantation site. For SIM at 30 and 60 days after implantation in bone defects, they were completely degraded and replaced by new bone with significantly superior morphology and mineral density, as compared with the control group.

Newly Formed Bone

All manuscripts concluded that simvastatin associated with calcium phosphate enhanced bone repair (Box 2). Although one article provided only descriptive histological analysis 2626. Adah F, Benghuzzi H, Tucci M, Russell G, England B. Effects of sustained release of statin by means of tricalcium phosphate lysine delivery system in defect and segmental femoral injuries on certain biochemical markers in vivo. Biomed Sci Instrum . 2006;42:126-35., seven studies showed improved bone formation with statistically significant results 1111. Santana WM de, Sousa DN de, Ferreira VM, Duarte WR. Simvastatin and biphasic calcium phosphate affects bone formation in critical-sized rat calvarial defects. Acta Cir Bras. 2016;31:300-307.,2727. Gao S, Shiota M, Fujii M, Chen K, Shimogishi M. Combination of simvastatin and hydroxyapatite fiber induces bone augmentation. Open J Regen Med 2013;2:53-60.,2828. Nyan M, Miyahara T, Noritake K, Hao J, Rodriguez R, Kuroda S, et al. Molecular and tissue responses in the healing of rat calvarial defects after local application of simvastatin combined with alpha tricalcium phosphate. J Biomed Mater Res Part B Appl Biomater 2010;93:65-73.,2929. Nyan M, Sato D, Kihara H, Machida T, Ohya K, Kasugai S. Effects of the combination with alpha-tricalcium phosphate and simvastatin on bone regeneration. Clin Oral Implants Res 2009;20:280-287.,3030. Rojbani H, Nyan M, Ohya K, Kasugai S. Evaluation of the osteoconductivity of α-tricalcium phosphate, β-tricalcium phosphate, and hydroxyapatite combined with or without simvastatin in rat calvarial defect. J Biomed Mater Res A 2011;98:488-498.,3131. Shahrezaie M, Moshiri A, Shekarchi B, Oryan A, Maffulli N, Parvizi J. Effectiveness of tissue engineered three-dimensional bioactive graft on bone healing and regeneration: an in vivo study with significant clinical value. J Tissue Eng Regen Med2017;1-15.,3232. Yin H, Li Y-G, Si M, Li J-M. Simvastatin-loaded macroporous calcium phosphate cement: preparation, in vitro characterization, and evaluation of in vivo performance. J Biomed Mater Res A 2012;100:2991-3000.. One article compared calcium phosphate alone and with SIM, without a control group (cloth) 2727. Gao S, Shiota M, Fujii M, Chen K, Shimogishi M. Combination of simvastatin and hydroxyapatite fiber induces bone augmentation. Open J Regen Med 2013;2:53-60., and seven studies showed that this association enhanced bone repair when compared with the control groups (cloth) 1111. Santana WM de, Sousa DN de, Ferreira VM, Duarte WR. Simvastatin and biphasic calcium phosphate affects bone formation in critical-sized rat calvarial defects. Acta Cir Bras. 2016;31:300-307.,2626. Adah F, Benghuzzi H, Tucci M, Russell G, England B. Effects of sustained release of statin by means of tricalcium phosphate lysine delivery system in defect and segmental femoral injuries on certain biochemical markers in vivo. Biomed Sci Instrum . 2006;42:126-35.,2828. Nyan M, Miyahara T, Noritake K, Hao J, Rodriguez R, Kuroda S, et al. Molecular and tissue responses in the healing of rat calvarial defects after local application of simvastatin combined with alpha tricalcium phosphate. J Biomed Mater Res Part B Appl Biomater 2010;93:65-73.,2929. Nyan M, Sato D, Kihara H, Machida T, Ohya K, Kasugai S. Effects of the combination with alpha-tricalcium phosphate and simvastatin on bone regeneration. Clin Oral Implants Res 2009;20:280-287.,3030. Rojbani H, Nyan M, Ohya K, Kasugai S. Evaluation of the osteoconductivity of α-tricalcium phosphate, β-tricalcium phosphate, and hydroxyapatite combined with or without simvastatin in rat calvarial defect. J Biomed Mater Res A 2011;98:488-498.,3131. Shahrezaie M, Moshiri A, Shekarchi B, Oryan A, Maffulli N, Parvizi J. Effectiveness of tissue engineered three-dimensional bioactive graft on bone healing and regeneration: an in vivo study with significant clinical value. J Tissue Eng Regen Med2017;1-15.,3232. Yin H, Li Y-G, Si M, Li J-M. Simvastatin-loaded macroporous calcium phosphate cement: preparation, in vitro characterization, and evaluation of in vivo performance. J Biomed Mater Res A 2012;100:2991-3000..

Box 2.
The effects of SIM carried by calcium phosphate on bone repair.

At two and eight weeks, 14 mg of tricalcium phosphate carrying 0.1 mg of SIM showed significantly higher bone volume than groups having 0, 0.01, 0.25, or 0.5 mg of SIM. In addition, the percentage of defect closure at eight weeks was significantly greater in the TCP-0.1 mg (97.86%±1.49) group than in the control group (41.67%±10.78) or the TCP-0.5 mg group (63.57%±9.3) 2929. Nyan M, Sato D, Kihara H, Machida T, Ohya K, Kasugai S. Effects of the combination with alpha-tricalcium phosphate and simvastatin on bone regeneration. Clin Oral Implants Res 2009;20:280-287.. However, another study using 40 mg HA fiber (HAF) showed that 0.45 mg of SIM induced significantly higher bone formation than lower amounts at four weeks and eight weeks. In this same study, the percentage of new bone volume (for each SIM/HAF mg ratio) was: 20.22%±5.53 (0/40 mg), 11.72%±3.53 (0.15/40 mg), 42.14%±4.43 (0.45/40 mg), and 31.22%±8.58 (0.75/40 mg), having significant differences between the 0/40 mg and the 0.45/40 mg groups (p=0.17) and between the 0.15/40 mg and the 0.45/40 mg groups (p=0.002) at eight weeks 2727. Gao S, Shiota M, Fujii M, Chen K, Shimogishi M. Combination of simvastatin and hydroxyapatite fiber induces bone augmentation. Open J Regen Med 2013;2:53-60..

According to Nyan et al, in the α-TCP group carrying 0.1 mg of SIM, most of the defects showed a complete bridging of new bone at the end of the experimental period due to a continuous regeneration of bone at the center. In addition, new bone was observed inside the remaining α-TCP particles 2929. Nyan M, Sato D, Kihara H, Machida T, Ohya K, Kasugai S. Effects of the combination with alpha-tricalcium phosphate and simvastatin on bone regeneration. Clin Oral Implants Res 2009;20:280-287.. In Nyan et al, a similar response was shown: 21 days after surgery, new bone deposition was observed in the middle of the α-TCP particles, which were almost completely surrounded by the regenerated bone 2828. Nyan M, Miyahara T, Noritake K, Hao J, Rodriguez R, Kuroda S, et al. Molecular and tissue responses in the healing of rat calvarial defects after local application of simvastatin combined with alpha tricalcium phosphate. J Biomed Mater Res Part B Appl Biomater 2010;93:65-73..

Most of the studies used TCP as biomaterial and presented positive results. Nyan et al. claimed that the percentage of new bone area was significantly higher in the α-TCP-SIM group (65.3%±6.3) as compared with the control group (15.8%±3.5) or the α-TCP-alone group (31.4%±3.8) at the end of experimental period (p<0.01) 2828. Nyan M, Miyahara T, Noritake K, Hao J, Rodriguez R, Kuroda S, et al. Molecular and tissue responses in the healing of rat calvarial defects after local application of simvastatin combined with alpha tricalcium phosphate. J Biomed Mater Res Part B Appl Biomater 2010;93:65-73.. Another study used β-TCP with 0.1 mg of SIM and showed in the SIM group a significantly greater amount of newly formed bone at six weeks (p<0.001) and at eight weeks (p<0.05) 3030. Rojbani H, Nyan M, Ohya K, Kasugai S. Evaluation of the osteoconductivity of α-tricalcium phosphate, β-tricalcium phosphate, and hydroxyapatite combined with or without simvastatin in rat calvarial defect. J Biomed Mater Res A 2011;98:488-498.. Furthermore, biphasic calcium phosphate (BCP) associated with SIM showed a higher area of newly formed bone tissue (12.71%±1.21) as compared with control group (0.00%±0.00) and BCP alone (9.04%±0.86) 1111. Santana WM de, Sousa DN de, Ferreira VM, Duarte WR. Simvastatin and biphasic calcium phosphate affects bone formation in critical-sized rat calvarial defects. Acta Cir Bras. 2016;31:300-307.. However, no significant results were observed using statins in association with HA particles 3030. Rojbani H, Nyan M, Ohya K, Kasugai S. Evaluation of the osteoconductivity of α-tricalcium phosphate, β-tricalcium phosphate, and hydroxyapatite combined with or without simvastatin in rat calvarial defect. J Biomed Mater Res A 2011;98:488-498.. Yin et al. 3232. Yin H, Li Y-G, Si M, Li J-M. Simvastatin-loaded macroporous calcium phosphate cement: preparation, in vitro characterization, and evaluation of in vivo performance. J Biomed Mater Res A 2012;100:2991-3000. used macroporous calcium phosphate cement, and through a histomorphometrical evaluation showed that the newly formed bone of the SIM group (7.4%±3.3) was significantly greater than in the non-SIM group (3.6%±1.4) (p<0.05).

Simvastatin (5 mg) in association with nanohydroxyapatite (nHA) had a synergistic effect, since the matrix of newly formed bone showed upper morphological characteristics compared with either nHA or SIM alone 3131. Shahrezaie M, Moshiri A, Shekarchi B, Oryan A, Maffulli N, Parvizi J. Effectiveness of tissue engineered three-dimensional bioactive graft on bone healing and regeneration: an in vivo study with significant clinical value. J Tissue Eng Regen Med2017;1-15.. Furthermore, another study also observed a positive effect in bone repair using 5 mg of SIM 2626. Adah F, Benghuzzi H, Tucci M, Russell G, England B. Effects of sustained release of statin by means of tricalcium phosphate lysine delivery system in defect and segmental femoral injuries on certain biochemical markers in vivo. Biomed Sci Instrum . 2006;42:126-35..

Discussion

Regarding bone regeneration, calcium phosphate as a statin carrier can be a valuable alternative to stem cells or growth factors. For that reason, this association (calcium phosphate and statin) can be recommended for clinical applications. This systematic review sought to provide information about how statins carried by calcium phosphate affect new bone formation and biocompatibility.

For bone healing, the effective dose of systemically administered simvastatin is higher than the routine dose in clinical applications (20-40 mg/day). For that reason, locally administered statin can be an alternative for achieving therapeutic concentrations for bone regeneration and avoiding the side effects of systemic doses 3333. Kheirallah M, Almeshaly H. Simvastatin , dosage and delivery system for supporting bone regeneration , an update review. J Oral Maxillofac Surgery, Med Pathol2016;28:205-209.. Given the possibility of a reduced inflammatory response with topical application, it is important to analyze the tissue response to locally administered statin. Shahrezaie et al. claimed that nHA and SIM improved the scaffold biocompatibility 3131. Shahrezaie M, Moshiri A, Shekarchi B, Oryan A, Maffulli N, Parvizi J. Effectiveness of tissue engineered three-dimensional bioactive graft on bone healing and regeneration: an in vivo study with significant clinical value. J Tissue Eng Regen Med2017;1-15., corroborating with another in vivo study which demonstrated that SIM significantly decreased pro-inflammatory cytokines and increased anti-inflammatory cytokine levels 3434. Jeong C, Kim SE, Shim K, Kim H, Song MH. Exploring the in vivo anti-inflammatory actions of simvastatin-loaded porous microspheres on inflamed tenocytes in a collagenase-induced animal model of achilles tendinitis. Int J Mol Sci 2018;19:820.. Although simvastatin presented topical anti-inflammatory properties, this response changes at higher doses. This fact can be demonstrated in Stein’s study, in which locally applied 2.2 mg of simvastatin caused significant inflammation in soft tissue in a rat mandible compared with lower doses 3535. Stein D, Lee Y, Schmid MJ, Killpack B, Genrich MA, Narayana N, et al. Local simvastatin effects on mandibular bone growth and inflammation. J Periodontol2005;76:1861-1870.. In addition, a large dose (10% wt) can produce an inflammatory response and induce a severe muscle necrosis around the biomaterial 3232. Yin H, Li Y-G, Si M, Li J-M. Simvastatin-loaded macroporous calcium phosphate cement: preparation, in vitro characterization, and evaluation of in vivo performance. J Biomed Mater Res A 2012;100:2991-3000.. Most of the manuscripts analyzed in this systematic review evaluated the inflammatory response and described no or a slight inflammatory reaction with low concentrations of SIM (0.1 to 0.45 mg) 1111. Santana WM de, Sousa DN de, Ferreira VM, Duarte WR. Simvastatin and biphasic calcium phosphate affects bone formation in critical-sized rat calvarial defects. Acta Cir Bras. 2016;31:300-307.,2727. Gao S, Shiota M, Fujii M, Chen K, Shimogishi M. Combination of simvastatin and hydroxyapatite fiber induces bone augmentation. Open J Regen Med 2013;2:53-60.,2828. Nyan M, Miyahara T, Noritake K, Hao J, Rodriguez R, Kuroda S, et al. Molecular and tissue responses in the healing of rat calvarial defects after local application of simvastatin combined with alpha tricalcium phosphate. J Biomed Mater Res Part B Appl Biomater 2010;93:65-73.,2929. Nyan M, Sato D, Kihara H, Machida T, Ohya K, Kasugai S. Effects of the combination with alpha-tricalcium phosphate and simvastatin on bone regeneration. Clin Oral Implants Res 2009;20:280-287.,3030. Rojbani H, Nyan M, Ohya K, Kasugai S. Evaluation of the osteoconductivity of α-tricalcium phosphate, β-tricalcium phosphate, and hydroxyapatite combined with or without simvastatin in rat calvarial defect. J Biomed Mater Res A 2011;98:488-498.,3232. Yin H, Li Y-G, Si M, Li J-M. Simvastatin-loaded macroporous calcium phosphate cement: preparation, in vitro characterization, and evaluation of in vivo performance. J Biomed Mater Res A 2012;100:2991-3000., which decreased over the experimental period in all studies.

In several studies, one of the main goals was to determine the optimal dose of simvastatin for inducing bone formation while minimizing the inflammatory response. Stein et al. stated that 0.1 mg of simvastatin in methylcellulose gel in a polylactic acid membrane resulted in minimal local inflammation, but it failed to stimulate significant bone growth 3535. Stein D, Lee Y, Schmid MJ, Killpack B, Genrich MA, Narayana N, et al. Local simvastatin effects on mandibular bone growth and inflammation. J Periodontol2005;76:1861-1870.. Nevertheless, when 0.1 mg of SIM (approximately 0.71% wt) was used with calcium phosphate carriers, it induced greater bone formation and caused only slight inflammation 1111. Santana WM de, Sousa DN de, Ferreira VM, Duarte WR. Simvastatin and biphasic calcium phosphate affects bone formation in critical-sized rat calvarial defects. Acta Cir Bras. 2016;31:300-307.,2828. Nyan M, Miyahara T, Noritake K, Hao J, Rodriguez R, Kuroda S, et al. Molecular and tissue responses in the healing of rat calvarial defects after local application of simvastatin combined with alpha tricalcium phosphate. J Biomed Mater Res Part B Appl Biomater 2010;93:65-73.,2929. Nyan M, Sato D, Kihara H, Machida T, Ohya K, Kasugai S. Effects of the combination with alpha-tricalcium phosphate and simvastatin on bone regeneration. Clin Oral Implants Res 2009;20:280-287.,3030. Rojbani H, Nyan M, Ohya K, Kasugai S. Evaluation of the osteoconductivity of α-tricalcium phosphate, β-tricalcium phosphate, and hydroxyapatite combined with or without simvastatin in rat calvarial defect. J Biomed Mater Res A 2011;98:488-498.. Furthermore, another study used different SIM doses (0, 0.15, 0.45, 0.75 mg of SIM in 40 mg of calcium phosphate), finding 0.45 mg of SIM to be the ideal concentration because it induced significantly greater bone formation than lower amounts but had no inflammatory response 2727. Gao S, Shiota M, Fujii M, Chen K, Shimogishi M. Combination of simvastatin and hydroxyapatite fiber induces bone augmentation. Open J Regen Med 2013;2:53-60.. Proportionally, 0.45 mg of SIM in 40 mg of carrier is nearly 1.1% wt; therefore, this study corroborates the findings of Yin et al, which showed that 1% wt SIM had no inflammatory reaction and improved bone formation 3232. Yin H, Li Y-G, Si M, Li J-M. Simvastatin-loaded macroporous calcium phosphate cement: preparation, in vitro characterization, and evaluation of in vivo performance. J Biomed Mater Res A 2012;100:2991-3000..

Several studies suggest that the anabolic bone effect of statins is dose-dependent at a certain level 2727. Gao S, Shiota M, Fujii M, Chen K, Shimogishi M. Combination of simvastatin and hydroxyapatite fiber induces bone augmentation. Open J Regen Med 2013;2:53-60.,2929. Nyan M, Sato D, Kihara H, Machida T, Ohya K, Kasugai S. Effects of the combination with alpha-tricalcium phosphate and simvastatin on bone regeneration. Clin Oral Implants Res 2009;20:280-287.,3636. Pauly S, Luttosch F, Morawski M, Haas NP, Schmidmaier G, Wildemann B. Simvastatin locally applied from a biodegradable coating of osteosynthetic implants improves fracture healing comparableto BMP-2 application Bone 2009;45:505-511., since higher doses showed more intense and prolonged inflammation. The inflammatory response under normal conditions is self-limited, contributing to the helping process by removing necrotic tissue, debris, and bacterial contaminants, as well as recruiting and activating fibroblasts 3737. Menke NB, Ward KR, Witten TM, Bonchev DG, Diegelmann RF Impaired woundhealing Clin Dermatol 2007;25:19-25.. However, higher doses may provoke an excessive inflammatory reaction, which could in turn stimulate the production of inflammatory cytokines and prevent the healing process 2727. Gao S, Shiota M, Fujii M, Chen K, Shimogishi M. Combination of simvastatin and hydroxyapatite fiber induces bone augmentation. Open J Regen Med 2013;2:53-60.,3737. Menke NB, Ward KR, Witten TM, Bonchev DG, Diegelmann RF Impaired woundhealing Clin Dermatol 2007;25:19-25..

SIM positively affects bone metabolism because it increases BMP-2 expression and suppresses osteoblast apoptosis, inducing osteoblastic differentiation, osteogenesis, and new bone formation. In addition, SIMs reduce bone loss, preventing osteoclast activity and osteoclastogenesis by decreasing the differentiation of macrophages or monocytes into osteoclast 3838. Oryan A, Kamali A, Moshiri A. Potential mechanisms and applications of statins on osteogenesis: Current modalities, conflicts and future directions. J Control Release 2015;215:12-24.,3939. Moshiri A, Sharifi AM, Oryan A. Role of Simvastatin on fracture healing and osteoporosis: a systematic review on in vivo investigations. Clin Exp Pharmacol Physiol2016;43:659-684.. Studies from this systematic review reinforce this information, since all manuscripts suggested that SIM associated with calcium phosphate enhanced bone regeneration, with seven out of eight studies producing statistically significant results 11,11. Santana WM de, Sousa DN de, Ferreira VM, Duarte WR. Simvastatin and biphasic calcium phosphate affects bone formation in critical-sized rat calvarial defects. Acta Cir Bras. 2016;31:300-307. 2727. Gao S, Shiota M, Fujii M, Chen K, Shimogishi M. Combination of simvastatin and hydroxyapatite fiber induces bone augmentation. Open J Regen Med 2013;2:53-60.,2828. Nyan M, Miyahara T, Noritake K, Hao J, Rodriguez R, Kuroda S, et al. Molecular and tissue responses in the healing of rat calvarial defects after local application of simvastatin combined with alpha tricalcium phosphate. J Biomed Mater Res Part B Appl Biomater 2010;93:65-73.,2929. Nyan M, Sato D, Kihara H, Machida T, Ohya K, Kasugai S. Effects of the combination with alpha-tricalcium phosphate and simvastatin on bone regeneration. Clin Oral Implants Res 2009;20:280-287.,3030. Rojbani H, Nyan M, Ohya K, Kasugai S. Evaluation of the osteoconductivity of α-tricalcium phosphate, β-tricalcium phosphate, and hydroxyapatite combined with or without simvastatin in rat calvarial defect. J Biomed Mater Res A 2011;98:488-498.,3131. Shahrezaie M, Moshiri A, Shekarchi B, Oryan A, Maffulli N, Parvizi J. Effectiveness of tissue engineered three-dimensional bioactive graft on bone healing and regeneration: an in vivo study with significant clinical value. J Tissue Eng Regen Med2017;1-15.,3232. Yin H, Li Y-G, Si M, Li J-M. Simvastatin-loaded macroporous calcium phosphate cement: preparation, in vitro characterization, and evaluation of in vivo performance. J Biomed Mater Res A 2012;100:2991-3000.. The positive effect of SIM was acknowledged when new bone formation area or volume significantly greater in the group that used SIM with calcium phosphate than the group of calcium phosphate alone. Therefore, this parameter was considered as a well succeed result.

It is a common sense that no animal model is capable of completely mimics human bone regarding macroscopy, microscopy and remodeling 4040. Pearce AI, Richards RG, Milz S, Schneider E, Pearce SG. Animal models for implant biomaterial research in bone: A review. European Cells and Materials 2007;13:1-10.. However, there are evidence that almost 50% of all animal models used to analyze bone repair are rats and rabbits 4141. Martini L, Fini M, Giavaresi G, Giardino R. Comparative medicine sheep model in orthopedic research: a literature review. Amer Assoc Lab Animal Sci2001;51:292-299.. Two different animal models were considered in this study, rat and rabbit. Nevertheless, the use of both models resulted in the same conclusion: calcium phosphate as a simvastatin carrier promoted significantly greater bone formation than the calcium phosphate alone. Indeed, the analysis of distinguished animal models reaching the same outcome reinforce the conclusion of this study. There are limitations in extrapolating results from animals to humans, but positive results in two distinct animal models can indicate more predictability of clinical outcomes. Several clinical studies already confirmed previous in vivo studies, showing a good agreement between pre-clinical and clinical studies regarding bone graft 4242. Wancket LM. Animal Models for Evaluation of bone implants and devices. Vet Pathol 2015;52:842-850.,4343. Li Y, Chen S-K, Li L, Qin L, Wang X-L, Lai Y-X. Bone defect animal models for testing efficacy of bone substitute biomaterials. J Orthop Transl2015;3:95-104..

Furthermore, in the included studies, the bone grafts were evaluated into two different types of bones: femur and calvaria, formed by distinct ossification process, endochondral and intramembranous, respectively. However, SIM significantly enhanced bone repair, despite the bone origin, reinforcing its positive effect.

Also, regarding the biomaterial preparation, it was observed that in most of the included studies, the incorporation of SIM to calcium phosphate was similar. First, SIM was dissolved in ethanol and then incorporated to calcium phosphate by dropping under sterile conditions or soaking. Only one study 3232. Yin H, Li Y-G, Si M, Li J-M. Simvastatin-loaded macroporous calcium phosphate cement: preparation, in vitro characterization, and evaluation of in vivo performance. J Biomed Mater Res A 2012;100:2991-3000. mixed SIM with the solid portion of the calcium phosphate cement, and then the solid and liquid phase were mixed. Independently of the biomaterial-SIM association process, the SIM group showed statistically higher new bone formation.

Another point to be analyzed is that one of the most important characteristics of biomaterial is biodegradability. In Shahrezaie’s study, the nHA, with or without SIM, degraded within 30 days after implantation and was replaced by woven bone 3131. Shahrezaie M, Moshiri A, Shekarchi B, Oryan A, Maffulli N, Parvizi J. Effectiveness of tissue engineered three-dimensional bioactive graft on bone healing and regeneration: an in vivo study with significant clinical value. J Tissue Eng Regen Med2017;1-15.. Furthermore, after 60 days they were replaced by corticomedullary compact bone. However, another study demonstrated that HA was the least degradable 3030. Rojbani H, Nyan M, Ohya K, Kasugai S. Evaluation of the osteoconductivity of α-tricalcium phosphate, β-tricalcium phosphate, and hydroxyapatite combined with or without simvastatin in rat calvarial defect. J Biomed Mater Res A 2011;98:488-498., showing that the nanostructure of the previous study may have contributed to its biodegradability by increasing the surface-to-volume ratio, which could, in turn, lead to a higher resorption rate 4444. Smolen D, Chudoba T, Malka I, Kedzierska A, Lojkowski W, Swieszkowski W, et al. Highly biocompatible, nanocrystalline hydroxyapatite synthesized in a solvothermal process driven by high energy density microwave radiation. Int J Nanomedicine2013;8:653-668.. Moreover, α-TCP and β-TCP are less stable than HA and are thus more soluble in aqueous environments 4545. Fernández E, Gil FJ, Ginebra MP, Driessens FC, Planell J a, Best SM. Calcium phosphate bone cements for clinical applications. Part I: solution chemistry. J Mater Sci Mater Med 1999;10:169-176., so β-TCP ceramics can be degraded more rapidly than HA ceramics in the body 4646. Fujita R, Yokoyama A, Kawasaki T, Kohgo T. Bone augmentation osteogenesis using hydroxyapatite and beta-tricalcium phosphate blocks. J Oral Maxillofac Surg. 2003;61:1045-53.,4747. Draenert M, Draenert A, Draenert K. Osseointegration of hydroxyapatite and remodeling-resorption of tricalciumphosphate ceramics. Microsc Res Tech 2013;76:370-380.. Furthermore, α-TCP was shown to have the most bioresorption, followed by β-TCP and HA 3030. Rojbani H, Nyan M, Ohya K, Kasugai S. Evaluation of the osteoconductivity of α-tricalcium phosphate, β-tricalcium phosphate, and hydroxyapatite combined with or without simvastatin in rat calvarial defect. J Biomed Mater Res A 2011;98:488-498.. In this same study, α-TCP and β-TCP carrying simvastatin exhibited more resorption, which can be explained by the degradation possibly being affected by the bone remodeling around the biomaterial 3030. Rojbani H, Nyan M, Ohya K, Kasugai S. Evaluation of the osteoconductivity of α-tricalcium phosphate, β-tricalcium phosphate, and hydroxyapatite combined with or without simvastatin in rat calvarial defect. J Biomed Mater Res A 2011;98:488-498..

Calcium phosphate biomaterials have been shown to be osteoconductive and biocompatible 33 . Suruagy AAP da S, Alves ATNN, Sartoretto SC, Calasans-Maia JA de A, Granjeiro JM, Calasans-Maia MD. Physico-chemical and histomorphometric evaluation of zinc-containing hydroxyapatite in rabbits calvaria. Braz Dent J2016;27:717-726.,4848. Ribeiro S, Sartoretto SC, Resende R, Uzeda M, Alves AT, Santos SA, et al. In vivo evaluation of Zinc-containing nanostructured carbonated hydroxyapatite. Key Engineering Materials2016;696:223-229.,4949. Resende RFB, Fernandes GVO, Santos SRA, Rossi AM, Lima I, Granjeiro JM, et al. Long-term biocompatibility evaluation of 0.5% zinc containing hydroxyapatite in rabbits. J Mater Sci Mater Med 2013 J;24:1455-1463.. Degradation of calcium phosphate may contribute to calcification due to its release of calcium and phosphate ions into the microenvironment, which could enhance bone formation 5050. Saito A, Suzuki Y, Kitamura M, Ogata S-I, Yoshihara Y, Masuda S, et al. Repair of 20-mm long rabbit radial bone defects using BMP-derived peptide combined with an alpha-tricalcium phosphate scaffold. Clin Exp Rheumatol 2006;77:700-706.. It could also work as a space maintainer, being highly advantageous in bone regeneration 2929. Nyan M, Sato D, Kihara H, Machida T, Ohya K, Kasugai S. Effects of the combination with alpha-tricalcium phosphate and simvastatin on bone regeneration. Clin Oral Implants Res 2009;20:280-287.. Calcium phosphate has been shown to be an ideal carrier for SIM because the association of SIM with biphasic calcium phosphate induced greater bone formation than SIM with collagen sponge 1111. Santana WM de, Sousa DN de, Ferreira VM, Duarte WR. Simvastatin and biphasic calcium phosphate affects bone formation in critical-sized rat calvarial defects. Acta Cir Bras. 2016;31:300-307.. In addition, the association of nHA with SIM showed a synergistic effect, inducing a greater bone volume compared with SIM or nHA alone 3131. Shahrezaie M, Moshiri A, Shekarchi B, Oryan A, Maffulli N, Parvizi J. Effectiveness of tissue engineered three-dimensional bioactive graft on bone healing and regeneration: an in vivo study with significant clinical value. J Tissue Eng Regen Med2017;1-15.. Furthermore, Stein et al. 3535. Stein D, Lee Y, Schmid MJ, Killpack B, Genrich MA, Narayana N, et al. Local simvastatin effects on mandibular bone growth and inflammation. J Periodontol2005;76:1861-1870.. Nevertheless, Nyan et al. used the same dosage of SIM carried by α-TCP and demonstrated a significantly higher bone formation when compared with the control group 2929. Nyan M, Sato D, Kihara H, Machida T, Ohya K, Kasugai S. Effects of the combination with alpha-tricalcium phosphate and simvastatin on bone regeneration. Clin Oral Implants Res 2009;20:280-287.. The difference may therefore be attributed to the simvastatin carrier. However, Rojbani et al. 3030. Rojbani H, Nyan M, Ohya K, Kasugai S. Evaluation of the osteoconductivity of α-tricalcium phosphate, β-tricalcium phosphate, and hydroxyapatite combined with or without simvastatin in rat calvarial defect. J Biomed Mater Res A 2011;98:488-498. demonstrated that the percentage of bone formation when SIM was carried by HA was not significant, suggesting that it could be associated with the slow biodegradability of HA.

In conclusion, the analyzed literature sustains that locally applied calcium phosphate as a SIM carrier is biocompatible and induces greater bone formation, which is statistically significant, regenerating from 18.3 to 107.97% more bone than calcium phosphate alone. The results so far reveal that calcium phosphate in association with SIM increases bone repair, being relevant for clinical regeneration of bone defects. Nevertheless, more standardized research is recommended since no optimum dose of the biomaterial has yet been defined because of the lack of a pattern in SIM dose and type of calcium phosphate.

Thus, the analysis of pertinent pre-clinical studies evidenced the calcium phosphate biocompatibility and its effectiveness in delivering SIM to improve the repair of bone defects. So, clinical trials are encouraged to investigate the impact of SIM associated with calcium phosphate bone graft in repairing bone defect in humans.

Acknowledgements

This work was supported by the Brazilian financial agencies CNPq (Grant no 457541/2013-0 and 467513/2014-7), FAPERJ(Grant no E-26/102.993/2012; E-26/203.012/2016), and Coordenação de Aperfeiçoamento de Pessoal de Nível Superior - Brasil (CAPES) (Finance Code 001), National Centre of Science and Technology on Regenerative Medicine - INCT Regenera (http://www.inctregenera.org.br/).

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Publication Dates

  • Publication in this collection
    12 June 2020
  • Date of issue
    Mar-Apr 2020

History

  • Received
    02 Aug 2019
  • Accepted
    11 Sept 2019
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