Reduction Of Osteogenic Differentiation Research Articles

Hypoxia-inducible Factor 1α Contributes to Matrix Metalloproteinases 2/9 and Inflammatory Responses in Periodontitis.

Periodontitis is a prevalent condition characterized by inflammation and tissue destruction within the periodontium, with hypoxia emerging as a contributing factor to its pathogenesis. Hypoxia-inducible factor 1α (HIF-1α) has a crucial role in orchestrating adaptive responses to hypoxic microenvironments and has been implicated in various inflammatory-related diseases. Understanding the interplay between HIF-1α, matrix metalloproteinases (MMPs), and inflammatory responses in periodontitis could provide insights into its molecular mechanisms. We investigated the relationship between HIF-1α, MMP2, and MMP9 in gingival crevicular fluid (GCF) and periodontal ligament stem cells (PDLSCs) from periodontitis patients. The expression levels of HIF-1α, MMP2, MMP9, and inflammatory factors (IL-6, IL-1β, TNF-α) were assessed using enzyme-linked immunosorbent assay (ELISA) and real-time PCR (RT-PCR). Additionally, osteogenic differentiation of PDLSCs was identified by alkaline phosphatase activity. Significantly elevated levels of HIF-1α, MMP2, and MMP9 were observed in GCF of periodontitis patients compared to controls. Positive correlations were found between HIF-1α and MMP2/MMP9, as well as with IL-6, IL-1β, and TNF-α. Modulation of HIF-1α expression in PDLSCs revealed its involvement in MMP2/9 secretion and inflammatory responses, with inhibition of HIF-1α mitigating these effects. Furthermore, HIF-1α inhibition alleviated the reduction in osteogenic differentiation induced by inflammatory stimuli. Our findings elucidate the regulatory role of HIF-1α in MMP expression, inflammatory responses, and osteogenic differentiation in periodontitis. In conclusion, targeting HIF-1α signaling pathways may offer therapeutic opportunities for managing periodontitis and promoting periodontal tissue regeneration.

Biological Function and Potential Applications of Garcinol in Human Dental Pulp Stem Cells

IntroductionThe regeneration of pulp tissue is crucial for true regenerative endodontic treatment, which requires a reduction in osteogenic differentiation. Garcinol, a histone acetyltransferase inhibitor, is a natural regulator that is known to suppress the osteogenic differentiation of dental pulp stem cells. In this study, the inhibitory effect of garcinol on the osteogenic differentiation of human dental pulp stem cells (hDPSCs) was evaluated using three-dimensional culture under in vitro and in vivo conditions. MethodshDPSCs were obtained from caries-free third molars and cultured with 10 μM garcinol for 7 days in an ultra-low attachment plate. The cell stemness and expression of osteogenic differentiation-related genes were analyzed using reverse transcription-polymerase chain reaction and single-cell analysis. A transplantation experiment was performed in mice to investigate whether garcinol-treated hDPSCs showed restrained osteogenic differentiation. ResultshDPSCs cultured in the U-shaped ultra-low attachment plate showed the highest expression of stemness-related genes. Garcinol-treated hDPSCs demonstrated downregulation of osteogenic differentiation, with lower expression of bone sialoprotein, which is related to bone formation, and higher expression of dentin sialophosphoprotein, which is related to dentin formation. However, the garcinol-treated hDPSCs did not show any alterations in their stemness. Consistent results were observed in the transplantation experiment in mice. ConclusionsGarcinol reduced the osteogenic differentiation of hDPSCs, which can contribute to true regenerative endodontic treatment.

Andrographis paniculata ameliorates estrogen deficiency-related osteoporosis by directing bone marrow mesenchymal stem cell fate.

Although Andrographis paniculata (AP) exhibits various biological functions such as anticancer, anti-inflammatory, antimalarial, antimicrobial, antioxidant, cardioprotective and immunomodulatory, its role in estrogen deficiency-related osteoporosis remains unclear. Ovariectomy (OVX)-induced estrogen deficiency-related osteoporotic mouse models and sham mouse models were established using 8-week-old female C57BL/6J mice. Micro-computed tomography (µCT) scanning was performed to assess the skeletal phenotype. The differentiation potential of bone marrow mesenchymal stem cells (BMSCs) from the OVX and sham groups was assessed by osteogenic or adipogenic induction medium in vitro. To verify the effects of AP, alizarin red S (ARS) staining, alkaline phosphatase (ALP) staining and oil red O (ORO) staining, reverse transcription assay and quantitative real-time polymerase chain reaction were applied to detect the lineage differentiation ability of BMSCs. µCT scanning showed that AP treatment attenuated the osteoporotic phenotype in OVX-induced estrogen deficiency-related osteoporotic mice. The results of ARS staining, ALP staining, ORO staining and quantitative real-time polymerase chain reaction indicated that BMSCs from OVX-induced osteoporotic mice displayed a significant reduction in osteogenic differentiation and an increase in adipogenic differentiation, which could be reversed by AP treatment. Our findings suggested that AP regulated the differentiation potential of BMSCs and ameliorated the development of estrogen deficiency-related osteoporosis, which might be an effective therapeutic method for estrogen deficiency-related osteoporosis.

Expression of microRNAs targeting heat shock protein B8 during in vitro expansion of dental pulp stem cells in regulating osteogenic differentiation

ObjectiveThe objectives of this study were (a) to determine the differentially expressed microRNAs that can target heat shock protein B8 (HspB8) during in vitro expansion of dental pulp stem cells (DPSCs); (b) to identify microRNAs involved in posttranscriptional regulation of HspB8 expression; and (c) to determine if HspB8-targeting microRNAs play roles on osteogenic differentiation of DPSCs. DesignDPSCs were established from rat first molars and expanded in vitro until the passage that cells lost osteogenic potential. TargetScan was used to predict the microRNAs that target HspB8 mRNA. Stem-loop quantitative RT-PCR was conducted to identify the HspB8-targeting microRNAs that were upregulated in late passages. The microRNAs mimics were transfected into DPSCs to assess their effects on HspB8 expression and on osteogenic differentiation. Resultslet-7b-5p, miR-98-5p, miR-215, miR-219a-1-3p and miR-295-5p were found to consistently increase expression in DPSCs after expansion. HspB8 mRNA and/or protein were significantly decreased in the DPSCs after transfection of miR-215 and miR-219a-1-3p mimics; whereas no significant reduction was seen after transfecting let-7b-5p, miR-98-5p and miR-295-5p mimics. When subjecting the transfected DPSCs to osteogenic induction, reduction of calcium deposition or osteogenic marker expression were observed with miR-215, miR-219a-1-3p and miR-295-5p transfection. ConclusionsIncreased expression of miR-215 and miR-219a-1-3p downregulates HspB8 expression, which contributes to the reduction of osteogenic capability of DPSCs. Increased expression of miR295-5p also causes a reduction of osteogenic differentiation, but not involved in HspB8.

Glycinol enhances osteogenic differentiation and attenuates the effects of age on mesenchymal stem cells.

Phytoestrogens, such as glycinol, have recently gained significant attention as an alternative therapy for osteoporosis due to their structural similarity to estradiol and their bone-generating potential. The osteogenic effects of glycinol were investigated in human bone marrow mesenchymal stem cells (BMSCs) derived from older (>50 years old) and younger subjects (<25 years old). BMSCs isolated from older donors demonstrated reduced osteogenesis. 17β-estradiol and glycinol exposure rescued the age-related reduction in osteogenic differentiation of BMSCs. These results correlated with the induction of osteogenic genes and estrogen receptor-α (ER-α) following glycinol treatment. ER antagonist studies further support that glycinol promotes osteogenesis through ER signaling. The results from these studies support investigating glycinol as a potential preventive or treatment for osteoporosis.

Cyclooxygenase-2 deficiency impairs muscle-derived stem cell-mediated bone regeneration via cellular autonomous and non-autonomous mechanisms.

This study investigated the role of cyclooxygenase-2 (COX-2) expression by donor and host cells in muscle-derived stem cell (MDSC)-mediated bone regeneration utilizing a critical size calvarial defect model. We found that BMP4/green fluorescent protein (GFP)-transduced MDSCs formed significantly less bone in COX-2 knock-out (Cox-2KO) than in COX-2 wild-type (WT) mice. BMP4/GFP-transduced Cox-2KO MDSCs also formed significantly less bone than transduced WT MDSCs when transplanted into calvarial defects created in CD-1 nude mice. The impaired bone regeneration in the Cox-2KO MDSCBMP4/GFP group is associated with downregulation of BMP4-pSMAD1/5 signaling, decreased osteogenic differentiation and lowered proliferation capacity after transplantation, compared with WT MDSCBMP4/GFP cells. The Cox-2KO MDSCBMP4/GFP group demonstrated a reduction in cell survival and direct osteogenic differentiation in vitro These effects were mediated in part by the downregulation of Igf1 and Igf2. In addition, the Cox-2KO MDSCBMP4/GFP cells recruited fewer macrophages than the WT MDSC/BMP4/GFP cells in the early phase after injury. We concluded that the bone regeneration capacity of Cox-2KO MDSCs was impaired because of a reduction in cell proliferation and survival capacities, reduction in osteogenic differentiation and a decrease in the ability of the cells to recruit host cells to the injury site.

Comparative study of osteogenic differentiation potential of mesenchymal stem cells derived from bone marrow and adipose tissue of osteoporotic female rats

Osteoporosis causes reduction of osteogenic differentiation of mesenchymal stem cells (MSCs) from bone marrow and adipose tissue. This study was designed to compare the osteogenic potential of bone marrow mesenchymal stem cells (BMMSCs) and adipose-derived stem cells (ADSCs) of ovariectomized (OVX) rats. MSC were harvested from bone marrow and inguinal fat pads of six OVX rats. The limitations of this report are that cells from different animals were pooled for the purpose of the experiments that were carried out in this study. At 7, 14 and 21 d of osteogenic differentiation, 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) conversion, alkaline phosphatase activity and gene expression for collagen I, osteocalcin, bone sialoprotein, osteopontin and bone morphogenetic protein-2 bone morphogenetic protein-2 (BMP-2) were analyzed. At 21 d, percentage of cells per field and percentage of mineralized nodule were analyzed. The data were subjected to analysis of variance, and the means were compared by Student–Newman–Keuls test. The cells, regardless of group, showed phenotypic characteristics consistent with stem cells. MTT conversion, alkaline phosphatase activity, percentage of mineralized nodule and expression of collagen I, osteocalcin and BMP-2 of ADSCs from OVX rats were higher when compared to BMMSCs from OVX rats in at least one of the evaluated periods (p < 0.05). However, bone sialoprotein and osteopontin expression were smaller than those observed in BMMSCs for all evaluated periods (p < 0.05). It was concluded that the ADSCs from OVX rats have higher osteogenic potential when compared to BMMSCs from OVX rats. This result suggests that the treatment of osteoporosis with autologous ADSCs may be more efficient.

Supplementation with Fish Oil and Genistein, Individually or in Combination, Protects Bone against the Adverse Effects of Methotrexate Chemotherapy in Rats

Cancer chemotherapy has been shown to induce long-term skeletal side effects such as osteoporosis and fractures; however, there are no preventative treatments. This study investigated the damaging effects of anti-metabolite methotrexate (MTX) subcutaneous injections (0.75 mg/kg BW) for five days and the potential protective benefits of daily oral gavage of fish oil at 0.5 mL/100 g BW (containing 375 mg of n-3 PUFA/100 g BW), genistein (2 mg/100 g BW), or their combination in young adult rats. MTX treatment alone significantly reduced primary spongiosa height and secondary spongiosa trabecular bone volume. Bone marrow stromal cells from the treated rats showed a significant reduction in osteogenic differentiation but an increase in adipogenesis ex vivo. Consistently, stromal cells had significantly higher mRNA levels of adipogenesis-related proliferator activator activated receptor-γ (PPAR-γ) and fatty acid binding protein (FABP4). MTX significantly increased the numbers of bone-resorbing osteoclasts and marrow osteoclast precursor cell pool while significantly enhancing the mRNA expression of receptor activator for nuclear factor kappa B ligand (RANKL), the RANKL/osteoprotegerin (OPG) ratio, interleukin-6 (IL-6), and tumor necrosis factor-α (TNF-α) in the bone. Supplementary treatment with fish oil and/or genistein significantly preserved trabecular bone volume and osteogenesis but suppressed MTX-induced adipogenesis and increases in osteoclast numbers and pro-osteoclastogenic cytokine expression. Thus, Fish oil and/or genistein supplementation during MTX treatment enabled not only preservation of osteogenic differentiation, osteoblast number and bone volume, but also prevention of MTX treatment-induced increases in bone marrow adiposity, osteoclastogenic cytokine expression and osteoclast formation, and thus bone loss.