Effects of drug discontinuation after short-term daily alendronate administration on osteoblasts and osteocytes in mice

In order to determine whether osteoclastic bone resorption is restarted after withdrawn of bisphosphonates, we conducted histological examinations on murine osteoclasts, osteoblasts and osteocytes after discontinuation of a daily regimen of alendronate (ALN) with a dosage of 1 mg/kg/day for 10 days. After drug discontinuation, metaphyseal trabecular number and bone volume remained unaltered for the first 4 days. Osteoclast number did not increase, while the number of apoptotic osteoclasts was elevated. On the other hand, tissue non-specific alkaline phosphatase-immunoreactive area was markedly reduced after ALN discontinuation. In addition, osteocytes showed an atrophic profile with empty lacunar areas during and after ALN treatment. Interestingly, as early as 36 h after a single ALN injection, osteocytes show signs of atrophy despite the presence of active osteoblasts. Structured illumination microscopy system showed shortening of osteocytic cytoplasmic processes after drug cessation, suggesting a possible morphological and functional disconnection between osteocytes and osteoblasts. Taken together, it appears that osteoclastic bone resorption is not resumed after ALN discontinuation; also, osteoblasts and osteocytes hardly seem to recover once they are inactivated and atrophied by ALN. In summary, it seems that one must pay more attention to the responses of osteoblasts and osteocytes, rather focusing on the resuming of osteoclastic bone resorption after the ALN discontinuation

A paradigm shift for bone quality in dentistry: A literature review

Purpose: The aim of this study was to present the current concept of bone quality based on the proposal by the National Institutes of Health (NIH) and some of the cellular and molecular factors that affect bone quality. Study selection: This is a literature review which focuses on collagen, biological apatite (BAp), and bone cells such as osteoblasts and osteocytes. Results: In dentistry, the term “bone quality” has long been considered to be synonymous with bone mineral density (BMD) based on radiographic and sensible evaluations. In 2000, the NIH proposed the concept of bone quality as “the sum of all characteristics of bone that influence the bone’s resistance to fracture,” which is completely independent of BMD. The NIH defines bone quality as comprising bone architecture, bone turnover, bone mineralization, and micro-damage accumulation. Moreover, our investigations have demonstrated that BAp, collagen, and bone cells such as osteoblasts and osteocytes play essential roles in controlling the current concept of bone quality in bone around hip and dental implants. Conclusion: The current concept of bone quality is crucial for understanding bone mechanical functions. BAp, collagen and osteocytes are the main factors affecting bone quality. Moreover, mechanical loading dynamically adapts bone quality. Understanding the current concept of bone quality is required in dentistry

Isotope microscopic assessment for localization of 15N-minodeonate in bone

Minodronate has been highlighted for its sustained effects on osteoporotic treatment. To determine the cellular mechanism of its sustained effects, we have assessed the localization of minodronate in mouse bones through isotope microscopy, by labeling it with a stable and rare nitrogen isotope (15N-minodronate). Eight-weeks-old male mice intravenously received 15N-minodronate (1 mg/kg) were fixed after three hours, 24 hrs, one week and one month. Isotope microscopy localized 15N-minodronate predominantly beneath osteoblasts (bone forming surface) rather than nearby osteoclasts (bone-resorbing surface). Literally, alendronate, another nitrogen-containing bisphosphonate, has been reported to accumulate on the bone-resorbing surface, and suddenly inhibit the osteoclasts. In contrast, minodronate appears to coat the bone-forming surface, without immediate inhibition of osteoclasts. A single injection of minodronate chronologically increased metaphyseal trabeculae, whereas the numbers of tartrate resistant acid phosphatase (TRAP)-positive osteoclasts and alkaline phosphatase (ALP)-reactive osteoblastic area were not reduced. Apoptotic osteoclasts were not apparent, but, finally being observed in the later stage of the experiments, while ALP-reactive osteoblasts were persisted on the trabeculae. Osteoclasts have developed ruffled borders at 3 hrs after minodronate administration; however, osteoclasts were roughly attached to the bone surfaces and did not form ruffled borders at 24 hrs after the administration. Von Kossa staining clearly demonstrated that osteoclasts did not incorporate the minodronate-treated bone matrix, while osteoclasts included abundant bone minerals inside in the control specimens. Taken together, minodronate accumulates in bone underneath osteoblasts rather than under bone-resorbing osteoclasts ; therefore, it is likely that the osteoclasts are not able to resorb and incorporate the minodronate-coated bone matrix, which may result in osteoclastic survival, avoiding osteoclastic apoptosis and consequently inducing cell coupling with osteoblasts. In conclusion, the resistance of miniodronate-coating bone from osteoclastic resorption, and the consequent cell coupling with osteoblasts appear to produce a long-lasting and bone-preserving effect

Clinical and imaging perspective and unanswered questions in a case of metronidazole induced encephalopathy

We discuss the clinical and imaging perspective in a case of a 78-year-old male who developed slurring of speech and ataxia acute in onset for the last 3 days. During his hospital stay, he developed multiple episodes of focal seizures without secondary generalization involving the angle of mouth on the right side. The patient had ataxia and positive cerebellar signs. In the past, the patient was treated for amoebic liver abscess and had undergone percutaneous aspiration of abscess. The patient was prescribed oral metronidazole and was discharged. This time, the patient underwent magnetic resonance imaging examination, which revealed lesion highly suggestive of metronidazole-induced encephalopathy. The offending drug was discontinued immediately after which the patient improved clinically. A follow-up scan was performed after 12 days and showed complete resolution of lesions

Bone-Orchestrating Cells, Osteocytes

Osteocytes build up functional syncytia, i.e., the osteocytic lacunar-canalicular system（OLCS）. The osteocytes are interconnected through gap junctions between their cytoplasmic processes, which pass through narrow passageways referred to as osteocytic canaliculi. There are two possible ways, in which molecules can be transported throughout the OLCS: via the cytoplasmic processes and their gap junctions, and via the pericellular space in the osteocytic canaliculi. Transport of minerals and small molecules through a spatially well-organized OLCS appears to be pivotal for bone mineral homeostasis and bone remodeling control. Recently, osteocyte-derived molecules -- sclerostin, dentin matrix protein-1, fibroblast growth factor 23（FGF23）-- have been put in evidence as they may be related to osteocytic functions such as regulation of bone remodeling and so forth. Osteocytes were shown to regulate phosphorus serum levels and osteoblastic activity through the expression of FGF23 and sclerostin. In our observations, FGF23 and sclerostin synthesis seemed to be associated with the spatial regularity of the OLCS: both proteins were consistently expressed by osteocytes in epiphyses and cortical bones showing regularly arranged OLCS. In contrast, mice bearing high bone turnover, e.g., osteoprotegerin deficient mice, revealed markedly-diminished sclerostin. This review will introduce our recent studies on the regularity of OLCS and the osteocytic function

Medial vascular calcification: a new concept challenging the classical paradigm of dystrophic calcification

Klotho deficient （kl/kl） mice are well known to　develop hyperphosphatemia and resultant Möncheberg’s vascular sclerosis, which consists of elongated or　fragmented elastic lamellae and abundant collagen fibrils inside the vessels. Instead of normal vascular smooth muscle cells （VSMCs）, the tunica media of the kl/kl aorta has cells rich with abundant endoplasmic reticulum and Golgi apparatus, somewhat resembling osteoblasts. There were many matrix vesicle-like structures and calcifying nodules in the vicinity of these osteoblast-like cells in kl/kl aorta. The calcifying nodules seem to trigger calcification in the elastic lamellae, without promoting it in the collagen fibrils inside the kl/kl aorta. Also, mineral deposition was observed within the intravascular amorphous organic component, suggesting dystrophic calcification. Thus, two possible pathways for vascular calcification exist: one mediated by matrix vesicle-like structures, and another taking place after the deposition of calcium phosphates in the amorphous organic component. Compared to the latter, which consists of the classical view of intravascular calcification, the former appears to mimic osteoblastic mineralization in bone, and could be the result of trans-differentiation of VSMCs into osteoblastic cells. In this work, we will review our current findings on the process of medial vascular calcification found in kl/kl mice

Immunolocalization of sclerostin synthesized by osteocytes in relation to bone remodeling in the interradicular septa of ovariectomized rats

This study aimed to elucidate whether estrogen deficiency would affect the synthesis of an osteocyte-derived factor, sclerostin, in the mesial region of alveolar bone. Eight 9-week-old Wistar female rats were ovariectomized (OVX) and the other eight rats were Sham-operated (Sham). After 4 weeks, the interradicular septa of mandibular first molar were embedded in paraffin, and then, were histochemically examined. Sclerostin-positive osteocytes were located in a superficial layer of the mesial region of Sham bone, while the OVX mesial region showed a lesser presence of the sclerostin-reactive osteocytes. There was no significant difference in the distribution of estrogen receptor α and TUNEL-positive cells in either Sham or OVX groups. Meanwhile, the Sham mesial region demonstrated many osteoclasts, but the OVX specimens showed numerous osteoclasts in association with intense immunolabeling of the receptor activator of the nuclear factor kB ligand. Complex meshwork of cement lines was seen consistent with irregularly-distributed osteocytic lacunar-canalicular system in the OVX mesial region, compared with those of the Sham specimens. In conclusion, estrogen deficiency appears to inhibit osteocytes for sclerostin synthesis in the mesial region of the interradicular septum, mediated by accelerated bone remodeling, rather than by directly effecting osteocytes

Altered distribution of bone matrix proteins and defective bone mineralization in klotho-deficient mice

In an attempt to identify the histological properties of the klotho-deficient (kl/kl) bone matrix, bone mineralization and the localization of Ca2+-binding bone matrix proteins - osteocalcin, dentin matrix protein-1 (DMP-1) and matrix Gla protein (MGP) - were examined in kl/kl tibiae. While a widespread osteocalcin staining could be verified in the wild-type bone matrix, localization of the same protein in the kl/kl tibiae seemed rather restricted to osteocytes with only a faint staining of the whole bone matrix. In wild-type mice, MGP immunoreactivity was present at the junction between the epiphyseal bone and cartilage, and at the insertion of the cruciate ligaments. In kl/kl mice, however, MGP was seen around the cartilaginous cores of the metaphyseal trabeculae and in the periphery of some cells of the bone surface. DMP-1 was identified in the osteocytic canalicular system of wild-type tibiae, but in the kl/kl tibiae this protein was mostly found in the osteocytic lacunae and in the periphery of some cells of the bone surface. Mineralization of the kl/kl bone seemed somewhat defective, with broad unmineralized areas within its matrix. In these areas, mineralized osteocytes along with their lacunae and osteocytic cytoplasmic processes were found to have intense osteocalcin and DMP-1 staining. Taken together, it might be that the excessive production of Ca2+-binding molecules such as osteocalcin and DMP-1 by osteocytes concentrates mineralization around such cells, disturbing the completeness of mineralization in the kl/kl bone matrix. (C) 2013 Elsevier Inc. All rights reserved