Eggshells as natural calcium carbonate source in combination with hyaluronan as beneficial additives for bone graft materials, an in vitro study

Introduction: In bone metabolism and the formation especially in bone substitution, calcium as basic module is of high importance. Different studies have shown that the use of eggshells as a bone substitute material is a promising and inexpensive alternative. In this in vitro study, the effects of eggshell granulate and calcium carbonate towards primary bovine osteoblasts were investigated. Hyaluronan (HA) was used as artificial extracellular matrix (ECM) for the used cells to facilitate proliferation and differentiation and to mimic the physiological requirements given by the egg in vivo. Methods: Hyaluronan, eggshells, a combination of hyaluronan and eggshells and CaCO3 were applied to the cells as additive to the used standard medium (modified High Growth Enhancement Medium) in a concentration of 0,1 g/l. The effect of the additives in the culture medium was examined by proliferation tests, immunohistochemical staining (anti-collagen type I, anti-osteopontin, anti-osteonectin and anti-osteocalcin) and kinetic oxygen measurements. Results: Our investigations revealed that all investigated additives show beneficial effect on osteoblast activity. Cell proliferation, differentiation and the metabolic activity of the differentiated cells could be influenced positively. Especially in the case cell cultures treated with eggshells the strongest effects were detected, while for the hyaluronan compared with eggshells, a weaker increase in cell activity was observed. Conclusion: In summary, it can be stated that the investigated components come into consideration as beneficial supplements for bone graft materials especially for maxillo facial surgery application.<br

Decrease in the expression of the type 1 PTH/PTHrP receptor (PTH1R) on chondrocytes in animals with osteoarthritis

<p>Abstract</p> <p>Background</p> <p>To evaluate the expression of the type 1 PTH/PTHrP receptor (PTH1R) on chondrocytes from hyaline cartilage over the course of osteoarthritis (OA).</p> <p>Methods</p> <p>In 12 NZW rabbits, the anterior cruciate ligament (ACL) was resected to create anterior instability of the knee. In 12 control rabbits, only a sham operation, without resection of the ACL, was performed. Four animals from each group were killed at 3, 6, and 12 weeks. After opening the knee joint, OA was macroscopically graded and hyaline cartilage of the load-bearing area was evaluated histologically according to the Mankin scale and by immunostaining for PTH1R.</p> <p>Results</p> <p>There was a positive linear correlation between the time after surgery and the macroscopic and histologic OA scores. The scores in the control group were constant over the time course. Immunostaining showed significantly less expression of PTH1R in the experimental compared to the control group after 6 (P < 0.05) and 12 weeks (P < 0.01). In the experimental group, a negative linear correlation between PTH1R expression and macroscopic and histologic grades was found.</p> <p>Conclusions</p> <p>The results show an in vivo decrease in the expression of PTH1R on chondrocytes over the time course of OA. Further studies are needed to evaluate whether new treatment approaches could evolve from this knowledge.</p

An in vitro study of osteoblast vitality influenced by the vitamins C and E

Vitamin C and vitamin E are known as important cellular antioxidants and are involved in several other non-antioxidant processes. Generally vitamin C and vitamin E are not synthesized by humans and therefore have to be applied by nutrition. The absence or deficiency of the vitamins can lead to several dysfunctions and even diseases (e.g. scurvy). The main interest in this study is that vitamin C and E are known to influence bone formation, e.g. vitamin C plays the key role in the synthesis of collagen, the major component of the extracellular bone matrix. In the present study we evaluate the effect of ascorbic acid (vitamin C) and α-tocopherol (vitamin E) on the proliferation and differentiation of primary bovine osteoblasts in vitro. Starting from standard growth medium we minimized the foetal calf serum to reduce their stimulatory effect on proliferation. An improved growth and an increased synthesis of the extracellular matrix proteins collagen type I, osteonectin and osteocalcin was observed while increasing the ascorbic acid concentration up to 200 μg/ml. Furthermore the effects of α-tocopherol on cell growth and cell differentiation were examined, whereby neither improved growth nor increased synthesis of the extracellular matrix proteins collagen type I, osteonectin and osteocalcin were detected. Further investigations are necessary to target at better supportive effect of vitamins on bone regeneration, and healing

Impact of testosterone on the expression of organic anion transporting polypeptides (OATP-1A2, OATP-2B1, OATP-3A1) in malignant and non-malignant human breast cells in vitro

Postmenopausal hormone therapy (HT) increases local estrogen formation in breast tissue. The enzymatic substrates depend on transmembrane anion transporting polypeptides (OATPs) to reach intracellular enzymes. The aim of this study was to investigate the effect of testosterone (T) on the expression of OATP-1A2, OATP-2B1, and OATP-3A1 in malignant (MCF-7, BT-474) and non-malignant (HBL-100) breast cells in vitro

In vitro inhibition of HUVECs by low dose methotrexate - Insights into oral adverse events

Background: With socio-economic changes, dentists and maxillofacial surgeons are more and more faced with medically compromised patients. Especially, the admission of antirheumatic drugs has increased remarkably. So dentists and maxillofacial surgeons should be aware of related adverse reactions that affect the craniofacial region. To identify possible cellular effects of disease modifying antirheumatic drugs (DMARDs) we investigated the influence of methotrexate (MTX) on human umbilical vein endothelial cells (HUVECs). Methods: HUVECs were incubated with various concentrations of MTX, corresponding to serum concentrations found in rheumatoid arthritis (RA) patients. The effect of MTX on cell proliferation, differentiation as well as mitochondrial activity was measured by use of immunostaining, cell counting and 3-(4, 5-dimethylthiazol-2-yl)- 2, 5-diphenyltetrazolium bromide (MTT) assay. Results: All samples incubated with MTX (1-1000 nM) showed significantly decreased cell viability when compared to controls. Cells were less proliferating, but did not lose their ability to synthesize endothelial proteins. A slight dose dependency of inhibiting effects was demonstrated. The observed differences between control and sample groups were rising with longer duration. Conclusion: Because of the crucial role of endothelial cells and their precursor cells in wound healing, a negative influence of MTX on oral health has to be supposed, correlating to clinical observations of adverse reactions in the oral cavity, such as ulcerative or erosive lesions

Short time administration of antirheumatic drugs - Methotrexate as a strong inhibitor of osteoblast's proliferation in vitro

<p>Abstract</p> <p>Introduction</p> <p>Due to increasing use of disease modifying antirheumatic drugs (DMARDs) as first line therapy in rheumatic diseases, dental and maxillofacial practitioner should be aware of drug related adverse events. Especially effects on bone-metabolism and its cells are discussed controversially. Therefore we investigate the in vitro effect of short time administration of low dose methotrexate (MTX) on osteoblasts as essential part of bone remodelling cells.</p> <p>Methods</p> <p>Primary bovine osteoblasts (OBs) were incubated with various concentrations of MTX, related to tissue concentrations, over a period of fourteen days by using a previously established standard protocol. The effect on cell proliferation as well as mitochondrial activity was assessed by using 3-(4, 5-dimethylthiazol-2-yl) 2, 5-diphenyltetrazolium bromide (MTT) assay, imaging and counting of living cells. Additionally, immunostaining of extracellular matrix proteins was used to survey osteogenic differentiation.</p> <p>Results</p> <p>All methods indicate a strong inhibition of osteoblast`s proliferation by short time administration of low dose MTX within therapeutically relevant concentrations of 1 to 1000nM, without affecting cell differentiation of middle-stage differentiated OBs in general. More over a significant decrease of cell numbers and mitochondrial activity was found at these MTX concentrations. The most sensitive method seems to be the MTT-assay. MTX-concentration of 0,01nM and concentrations below had no inhibitory effects anymore.</p> <p>Conclusion</p> <p>Even low dose methotrexate acts as a potent inhibitor of osteoblast’s proliferation and mitochondrial metabolism in vitro, without affecting main differentiation of pre-differentiated osteoblasts. These results suggest possible negative effects of DMARDs concerning bone healing and for example osseointegration of dental implants. Especially the specifics of the jaw bone with its high vascularisation and physiological high tissue metabolism, suggests possible negative effects of DMARD therapy concerning oral and cranio-maxillofacial bone surgery as could be seen in a similar way in bisphosphonate related osteonecrosis of the jaw.</p