Cell and Tissue Research

, Volume 361, Issue 2, pp 467–476 | Cite as

Effects of the 1, 4-dihydropyridine L-type calcium channel blocker benidipine on bone marrow stromal cells

  • Zhong-ping Ma
  • Jia-cheng Liao
  • Chang Zhao
  • Dao-zhang CaiEmail author
Regular Article


Osteoporosis (OP) often increases the risk of bone fracture and other complications and is a major clinical problem. Previous studies have found that high blood pressure is associated with bone formation abnormalities, resulting in increased calcium loss. We have investigated the effect of the antihypertensive drug benidipine on bone marrow stromal cell (BMSC) differentiation into osteoblasts and bone formation under osteoporotic conditions. We used a combination of in vitro and in vivo approaches to test the hypothesis that benidipine promotes murine BMSC differentiation into osteoblasts. Alkaline phosphatase (ALP), osteocalcin (OCN), runt-related transcription factor 2 (RUNX2), β-catenin, and low-density lipoprotein receptor-related protein 5 (LRP5) protein expression was evaluated in primary femoral BMSCs from C57/BL6 mice cultured under osteogenic conditions for 2 weeks to examine the effects of benidipine. An ovariectomized (OVX) mouse model was used to investigate the effect of benidipine treatment for 3 months in vivo. We found that ALP, OCN, and RUNX2 expression was up-regulated and WNT/β-catenin signaling was enhanced in vitro and in vivo. In OVX mice that were intragastrically administered benidipine, bone parameters (trabecular thickness, bone mineral density, and trabecular number) in the distal femoral metaphysis were significantly increased compared with control OVX mice. Consistently, benidipine promoted BMSC differentiation into osteoblasts and protected against bone loss in OVX mice. Therefore, benidipine might be a suitable candidate for the treatment of patients with postmenopausal osteoporosis and hypertension.


Osteoblast Benidipine WNT signal pathway Bone metabolism Osteoporosis 



We thank Hua Wu (Hohhot Zhongke Medical Technology, Inner Mongolia, PR China) for excellent technical support with micro-CT.

Supplementary material

441_2015_2115_Fig7_ESM.gif (29 kb)
Supplementary Fig. 1

BD is not able to promote changes in osteogenic markers such as OCN (b) and RUNX2 (c) in the absence of differentiation medium for 2 weeks. Columns represent the means ± SD from three wells per group (b, c). (GIF 29 kb)

441_2015_2115_MOESM1_ESM.tif (830 kb)
High Resolution Image (TIFF 829 kb)
441_2015_2115_Fig8_ESM.gif (218 kb)
Supplementary Fig. 2

BD has no effect on endogenous stem cells. As shown by the results of immunohistochemistry with the characteristic markers of marrow stromal cells, such as CD105 (a) and CD106 (b), endogenous stem cells do not change significantly after BD treatment. CD105- (c) and CD106- (d) positive cells are evenly distributed in all groups as demonstrated microscopically by immunohistochemical staining, with no significant differences between each other. Columns represent the means ± SD from six mice per group (c, d). (GIF 218 kb)

441_2015_2115_MOESM2_ESM.tif (9.3 mb)
High Resolution Image (TIFF 9477 kb)


  1. Baron R, Rawadi G (2007) Targeting the Wnt/beta-catenin pathway to regulate bone formation in the adult skeleton. Endocrinology 148:2635–2643PubMedCrossRefGoogle Scholar
  2. Bianco P, Kuznetsov SA, Riminucci M, Gehron Robey P (2006) Postnatal skeletal stem cells. Methods Enzymol 419:117–148PubMedCrossRefGoogle Scholar
  3. Boyden LM, Mao J, Belsky J, Mitzner L, Farhi A, Mitnick MA, Dianqing W, Insoga K, Lifton RP (2002) High bone density due to a mutation in LDL-receptor–related protein 5. Radiology 346:1513–1521Google Scholar
  4. Brickman AS, Nyby MD, Hungen K von, Eggena P, Tuck ML (1990) Calcitropic hormones, platelet calcium, and blood pressure in essential hypertension. Hypertension 16:515–522Google Scholar
  5. Canalis E, Giustina A, Bilezikian JP (2007) Mechanisms of anabolic therapies for osteoporosis. N Engl J Med 357:905–916PubMedCrossRefGoogle Scholar
  6. Cappuccio FP, Meilahn E, Zmuda JM, Cauley JA (1999) High blood pressure and bone-mineral loss in elderly white women: a prospective study. Study of Osteoporotic Fractures Research Group. Lancet 354:971–975PubMedCrossRefGoogle Scholar
  7. Chan GK, Duque G (2002) Age-related bone loss: old bone, new facts. Gerontology 48:62–71PubMedCrossRefGoogle Scholar
  8. Cirillo M, Galletti F, Strazzullo P, Torielli L, Melloni MC (1989) On the pathogenetic mechanism of hypercalciuria in genetically hypertensive rats of the Milan strain. Am J Hypertens 2:741–746PubMedCrossRefGoogle Scholar
  9. Danks L, Takayanagi H (2013) Immunology and bone. J Biochem 154:29–39PubMedCrossRefGoogle Scholar
  10. Davey RA, Clarke MV, Sastra S, Skinner JP, Chiang C, Anderson PH, Zajac JD (2012) Decreased body weight in young Osterix-Cre transgenic mice results in delayed cortical bone expansion and accrual. Transgenic Res 21:885–893PubMedCrossRefGoogle Scholar
  11. Gadallah M, Massry SG, Bigazzi R, Horst RL, Eggena P, Campese VM (1991) Intestinal absorption of calcium and calcium metabolism in patients with essential hypertension and normal renal function. Am J Hypertens 4:404–409PubMedCrossRefGoogle Scholar
  12. Gong Y, Slee RB, Fukai N, Rawadi G, Roman-Roman S, Reginato AM, Wang H, Cundy T, Glorieux FH, Lev D, Zacharin M, Oexle K, Marcelino J, Suwairi W, Heeger S, Sabatakos G, Apte S, Adkins WN, Allgrove J, Arslan-Kirchner M, Batch JA, Beighton P, Black GC, Boles RG, Boon LM, Borrone C, Brunner HG, Carle GF, Dallapiccola B, De Paepe A, Floege B, Halfhide ML, Hall B, Hennekam RC, Hirose T, Jans A, Jüppner H, Kim CA, Keppler-Noreuil K, Kohlschuetter A, LaCombe D, Lambert M, Lemyre E, Letteboer T, Peltonen L, Ramesar RS, Romanengo M, Somer H, Steichen-Gersdorf E, Steinmann B, Sullivan B, Superti-Furga A, Swoboda W, Boogaard MJ van den, Van Hul W, Vikkula M, Votruba M, Zabel B, Garcia T, Baron R, Olsen BR, Warman ML, Osteoporosis-Pseudoglioma Syndrome Collaborative Group (2001) LDL receptor-related protein 5 (LRP5) affects bone accrual and eye development. Cell 107:513–523Google Scholar
  13. Grobbee DE, Hackeng WH, Birkenhager JC, Hofman A (1988) Raised plasma intact parathyroid hormone concentrations in young people with mildly raised blood pressure. Br Med J (Clin Res Ed) 296:814–816CrossRefGoogle Scholar
  14. Hu H, Hilton MJ, Tu X, Yu K, Ornitz DM, Long F (2005) Sequential roles of Hedgehog and Wnt signaling in osteoblast development. Development 132:49–60PubMedCrossRefGoogle Scholar
  15. Hvarfner A, Bergstrom R, Morlin C, Wide L, Ljunghall S (1987) Relationships between calcium metabolic indices and blood pressure in patients with essential hypertension as compared with a healthy population. J Hypertens 5:451–456PubMedCrossRefGoogle Scholar
  16. Izawa Y, Sagara K, Kadota T, Makita T (1985) Bone disorders in spontaneously hypertensive rat. Calcif Tissue Int 37:605–607PubMedCrossRefGoogle Scholar
  17. Karsenty G (2003) The complexities of skeletal biology. Nature 423:316–318PubMedCrossRefGoogle Scholar
  18. Katz AM, Hager WD, Messineo FC, Pappano AJ (1984) Cellular actions and pharmacology of the calcium channel blocking drugs. Am J Med 77:2–10PubMedCrossRefGoogle Scholar
  19. Kosaka N, Uchii M (1998) Effect of benidipine hydrochloride, a dihydropyridine-type calcium antagonist, on the function of mouse osteoblastic cells. Calcif Tissue Int 62:554–556PubMedCrossRefGoogle Scholar
  20. Kronenberg HM (2003) Developmental regulation of the growth plate. Nature 423:332–336PubMedCrossRefGoogle Scholar
  21. Little RD, Carulli JP, Del Mastro RG, Dupuis J, Osborne M, Folz C, Manning SP, Swain PM, Zhao SC, Eustace B, Lappe MM, Spitzer L, Zweier S, Braunschweiger K, Benchekroun Y, Hu X, Adair R, Chee L, FitzGerald MG, Tulig C, Caruso A, Tzellas N, Bawa A, Franklin B, McGuire S, Nogues X, Gong G, Allen KM, Anisowicz A, Morales AJ, Lomedico PT, Recker SM, Van Eerdewegh P, Recker RR, Johnson ML (2002) A mutation in the LDL receptor-related protein 5 gene results in the autosomal dominant high-bone-mass trait. Am J Hum Genet 70:11–19PubMedCentralPubMedCrossRefGoogle Scholar
  22. McCarron DA, Pingree PA, Rubin RJ, Gaucher SM, Molitch M, Krutzik S (1980) Enhanced parathyroid function in essential hypertension: a homeostatic response to a urinary calcium leak. Hypertension 2:162–168PubMedCrossRefGoogle Scholar
  23. Melton LJ 3rd, Chrischilles EA, Cooper C, Lane AW, Riggs BL (1992) Perspective. How many women have osteoporosis? J Bone Miner Res 7:1005–1010PubMedCrossRefGoogle Scholar
  24. Nishiya Y, Sugimoto S (2001) Effects of various antihypertensive drugs on the function of osteoblast. Biol Pharm Bull 24:628–633PubMedCrossRefGoogle Scholar
  25. Nishiya Y, Kosaka N, Uchii M, Sugimoto S (2002) A potent 1,4-dihydropyridine L-type calcium channel blocker, benidipine, promotes osteoblast differentiation. Calcif Tissue Int 70:30–39PubMedCrossRefGoogle Scholar
  26. Owen M (1988) Marrow stromal stem cells. J Cell Sci Suppl 10:63–76PubMedCrossRefGoogle Scholar
  27. Rodda SJ, McMahon AP (2006) Distinct roles for Hedgehog and canonical Wnt signaling in specification, differentiation and maintenance of osteoblast progenitors. Development 133:3231–3244PubMedCrossRefGoogle Scholar
  28. Rodrıguez JP, Garat S, Gajardo H, Pino AM, Seitz G (1999) Abnormal osteogenesis in osteoporotic patients is reflected by altered mesenchymal stem cells dynamics. J Cell Biochem 75:414–423PubMedCrossRefGoogle Scholar
  29. Rossini M, Gatti D, Adami S (2013) Involvement of WNT/beta-catenin signaling in the treatment of osteoporosis. Calcif Tissue Int 93:121–132PubMedCrossRefGoogle Scholar
  30. Strazzullo P, Nunziata V, Cirillo M, Giannattasio R, Ferrara LA, Mattioli PL, Mancini M (1983) Abnormalities of calcium metabolism in essential hypertension. Clin Sci 65:137–141PubMedGoogle Scholar
  31. Sun H, Kim JK, Mortensen R, Mutyaba LP, Hankenson KD, Krebsbach PH (2013) Osteoblast-targeted suppression of PPARgamma increases osteogenesis through activation of mTOR signaling. Stem Cells 31:2183–2192PubMedCrossRefGoogle Scholar
  32. Wang J, Bi M, Zhu Z, Wu L, Wang J (2014) Effects of the antihypertensive drug benidipine on osteoblast function in vitro. Exp Ther Med 7:649–653PubMedCentralPubMedGoogle Scholar
  33. Young EW, Morris CD, McCarron DA (1992) Urinary calcium excretion in essential hypertension. J Lab Clin Med 120:624–632PubMedGoogle Scholar

Copyright information

© Springer-Verlag Berlin Heidelberg 2015

Authors and Affiliations

  • Zhong-ping Ma
    • 1
    • 2
    • 3
  • Jia-cheng Liao
    • 1
    • 2
    • 4
  • Chang Zhao
    • 1
    • 2
  • Dao-zhang Cai
    • 1
    • 2
    Email author
  1. 1.Department of OrthopedicsThe Third Affiliated Hospital of Southern Medical UniversityGuangzhouPeople’s Republic of China
  2. 2.Academy of OrthopedicsGuangzhouPeople’s Republic of China
  3. 3.Department of OrthopedicsThe Second Affiliated Hospital of Inner Mongolia Medical UniversityHohhotPeople’s Republic of China
  4. 4.Department of Orthopedic SurgeryThe Affiliated Longhua People’s Hospital of Southern Medical UniversityShenzhenPeople’s Republic of China

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