Basic Study
Copyright ©The Author(s) 2020. Published by Baishideng Publishing Group Inc. All rights reserved.
World J Exp Med. Jan 8, 2020; 10(1): 1-9
Published online Jan 8, 2020. doi: 10.5493/wjem.v10.i1.1
Alendronate disturbs femoral growth due to changes during immunolocalization of transforming growth factor-β1 and bone morphogenetic protein-2 in epiphyseal plate
Juliana Souza Vieira, Emanuelle Juliana Cunha, Juliana Feltrin de Souza, Luis Henrique Koeler Chaves, Jessica Lakes de Souza, Allan Fernando Giovanini
Juliana Souza Vieira, Emanuelle Juliana Cunha, Luis Henrique Koeler Chaves, Jessica Lakes de Souza, Allan Fernando Giovanini, Clinical Dentistry, University Positivo, Curitiba, PR 81280-330, Brazil
Juliana Feltrin de Souza, Stomatology Department, University Federal of Paraná, Curitiba, PR 80210-170, Brazil
Author contributions: Vieira JS and Cunha EJ performed equally the majority of laboratorial experiments; de Souza JF, Chaves LHK and de Souza JL participated in treatment of animals and performed the immunohistochemical investigations; Giovanini AF analyzed all data and wrote the paper.
Institutional animal care and use committee statement: The principles of laboratory animal care and national laws on animal use were observed for the present study with the approval of the Ethical Committee for Animal Research of Positive University (Protocol #266).
Conflict-of-interest statement: Nothing to disclose.
Data sharing statement: No additional data are available.
ARRIVE guidelines statement: The authors have read the ARRIVE guidelines, and the manuscript was prepared and revised according to the ARRIVE guidelines.
Open-Access: This article is an open-access article which was selected by an in-house editor and fully peer-reviewed by external reviewers. It is distributed in accordance with the Creative Commons Attribution Non Commercial (CC BY-NC 4.0) license, which permits others to distribute, remix, adapt, build upon this work non-commercially, and license their derivative works on different terms, provided the original work is properly cited and the use is non-commercial. See:
Corresponding author: Allan Fernando Giovanini, MD, PhD, Associate Professor, Clinical Dentistry, University Positivo, R Pedro Viriato Parigot de Souza, 5300 Campo Comprido, Curitiba, PR 81280-330, Brazil.
Received: May 6, 2019
Peer-review started: May 6, 2019
First decision: September 2, 2019
Revised: November 26, 2019
Accepted: December 14, 2019
Article in press: December 14, 2019
Published online: January 8, 2020

The epiphyseal growth plate is an important anatomical segment localized on the ends of a long bone. Despite the abovementioned atractive reasons for alendronate’s use, few data on the effect of alendronate during epiphyseal growth exist.


Verify the effect of alendronate on the growth epiphyseal plate, and compare its effect with the size of the femur during the double-staining of the immunolocalization of transforming growth factor-β1 (TGF-β1) and bone morphogenetic protein-2 (BMP2) in endochondral ossifing in specimens that have received alendronate.


Forty newborn rats were randomly divided into two groups: a control group (were given applications of 1 mg/kg physiologic saline) and a group that received Alendronate (a dose of 2.5 mg/kg). These groups were then divided into two subgroups for euthanasia in two and 12 d of life. After euthanasia, the femurs were removed, and the femoral bones were measured linearly between the apex of the greater trochanter until the lower intercondylar midlle face to verify the probable bone growth between 3 and 12 d in control and alednroanto treated rats. Posteriorly, the surgical pieces were also sent to the histopathology laboratory to produce histological slides. The obtained slides were stained with hematoxylin and eosin to measure each of the cartilage zones in endochondral development. and other slides were immunohistochemically tested for anti- TGF-β1 and BMP-2 antibodies to investigate the immunolocalization of these proteins in the epiphyseal plaque area.


On the third day, some diferences between the control group and specimens treated with alendronate were verified. Macroscopiccaly, we found similarities in size between the femoral bones when we compared the control group with the specimens that received alendronate. On the 12th day, the bone size of the mice receiving the drug was significantly smaller than those of the control group. These results coincide with changes in the TGF-β1 and BMP-2 expression. In the specimens that received alendronate, the TGF-β1 was expressed in some sites of trabecular bone that was neoformed, peripherally to the bone marrow area. The BMP-2 was also positive in proliferative chondrocytes and hypertrofic chondrocytes. On the 12th day, all layers of chondrocytes exhibited positivity for BMP-2 in the specimens that received alendronate. In the interface between the trabecular bone and cartilage, an area of disorganized bone deposition was evident. Neoformed bone also appeared to be different at 12 d. In the control group, BMP-2 was positive in an intense area of bone trabeculae, whereas the alendronate-treated group showed TGF-β1 positive trabeculae and a greater bone area.


Alendronate alters the immunolocalization of TGF-β1 and BMP-2 simultaneously, a condition that changes the usual histological aspects of the cartilage zone and impairs epiphysis growth and femur growth.

Keywords: Alendronate, Bone development, Epiphyseal plate, Bone morphogentic protein-2, Transforming growth factor-β1

Core tip: This is the first study that addresses the likely action of alendronate in appendicular bone development. Through of macroscopic, immunohistochemical and histological analysis, we revealed that alendronatemay impairs the bone growth, since alters the usual development of epiphyseal plate associated to changes in the immunolocalization of transforming growth factor-β1 and bone morphogenetic protein-2.