Genome-Wide CRISPR/Cas9-Based Screening for Deubiquitinase Subfamily Identifies Ubiquitin-Specific Protease 11 as a Novel Regulator of Osteogenic Differentiation

The osteoblast differentiation capacity of mesenchymal stem cells must be tightly regulated, as inadequate bone mineralization can lead to osteoporosis, and excess bone formation can cause the heterotopic ossification of soft tissues. The balanced protein level of Msh homeobox 1 (MSX1) is critical during normal osteogenesis. To understand the factors that prevent MSX1 protein degradation, the identification of deubiquitinating enzymes (DUBs) for MSX1 is essential. In this study, we performed loss-of-function-based screening for DUBs regulating MSX1 protein levels using the CRISPR/Cas9 system. We identified ubiquitin-specific protease 11 (USP11) as a protein regulator of MSX1 and further demonstrated that USP11 interacts and prevents MSX1 protein degradation by its deubiquitinating activity. Overexpression of USP11 enhanced the expression of several osteogenic transcriptional factors in human mesenchymal stem cells (hMSCs). Additionally, differentiation studies revealed reduced calcification and alkaline phosphatase activity in USP11-depleted cells, while overexpression of USP11 enhanced the differentiation potential of hMSCs. These results indicate the novel role of USP11 during osteogenic differentiation and suggest USP11 as a potential target for bone regeneration.

BMP3 Affects Cortical and Trabecular Long Bone Development in Mice

Bone morphogenetic proteins (BMPs) have a major role in tissue development. BMP3 is synthesized in osteocytes and mature osteoblasts and has an antagonistic effect on other BMPs in bone tissue. The main aim of this study was to fully characterize cortical bone and trabecular bone of long bones in both male and female Bmp3−/− mice. To investigate the effect of Bmp3 from birth to maturity, we compared Bmp3−/− mice with wild-type littermates at the following stages of postnatal development: 1 day (P0), 2 weeks (P14), 8 weeks and 16 weeks of age. Bmp3 deletion was confirmed using X-gal staining in P0 animals. Cartilage and bone tissue were examined in P14 animals using Alcian Blue/Alizarin Red staining. Detailed long bone analysis was performed in 8-week-old and 16-week-old animals using micro-CT. The Bmp3 reporter signal was localized in bone tissue, hair follicles, and lungs. Bone mineralization at 2 weeks of age was increased in long bones of Bmp3−/− mice. Bmp3 deletion was shown to affect the skeleton until adulthood, where increased cortical and trabecular bone parameters were found in young and adult mice of both sexes, while delayed mineralization of the epiphyseal growth plate was found in adult Bmp3−/− mice.

Elaborate the Mechanism of Ancient Classic Prescriptions (Erzhi Formula) in Reversing GIOP by Network Pharmacology Coupled with Zebrafish Verification

Osteoporosis is a degenerative disease that endangers human health. At present, chemical drugs used for osteoporosis have serious side effects. Therefore, it is valuable to search herbs with high safety and good curative effect in antiosteoporosis. Erzhi formula (EZF), an ancient classic compound, has been reported to have a beneficial effect in antiosteoporosis, but its mechanism is unclear. In this paper, the active compounds of EZF were found in Systems Pharmacology Database, and gene targets related to osteoporosis were obtained in GeneCards. The GO functional and KEGG pathway enrichment analysis were performed by Metascape. The network of “components-targets-signal pathway” was constructed by Cytoscape. Next, molecular docking between the active components and hub genes related to the PI3K-Akt signaling pathway was conducted by Autodock. In the verification experiment, the zebrafish induced by prednisolone (PNSL) was used to reproduce glucocorticoid-induced osteoporosis (GIOP) model, and then the reversal effects of EZF were systematically evaluated according to the behavior, skull staining area, bone mineralization area (BMA), average optical density (AOD), and cumulative optical density (COD). Finally, it was shown that 24 components in EZF could regulate 39 common gene targets to exert antiosteoporosis effect. Besides, the main regulatory mechanisms of EZF were 4 signaling pathways: PI3K-Akt, JAK-STAT, AGE-RAGE, and cancer pathway. In PI3K-Akt signaling pathway, wedelolactone, dimethyl wedelolactone, specnuezhenide, ursolic acid, acacetin, beta-sitosterol, apigenin, and kaempferol can bind tightly with EGF, IL-2, and IL-4 genes. Compared with the model group, the moving distance, swimming speed, and cumulative swimming time of zebrafish in EZF group were significantly increased ( P < 0.05 ). Meanwhile, the BMA and COD of zebrafish were significantly improved after the intervention of EZF ( P < 0.05 ). In summary, the 24 components of EZF exert their antiosteoporosis effects by regulating 39 related gene targets, among which the PI3K signaling pathway is crucial. EZF can promote bone formation and reversed GIOP through “multicomponent/multitarget/multipathway” and the medium dose of EZF may be the most suitable concentration for the treatment of GIOP in zebrafish model.

The Impact of Vitamin K2 (Menaquionones) in Children’s Health and Diseases: A Review of the Literature

Vitamin K2 activates vitamin K-dependent proteins that support many biological functions, such as bone mineralization, the inhibition of vascular stiffness, the improvement of endothelial function, the maintenance of strong teeth, brain development, joint health, and optimal body weight. Due to the transformation of food habits in developed countries over the last five decades, vitamin K and, specifically, vitamin K2 intakes among parents and their offspring have decreased significantly, resulting in serious health implications. The therapeutics used in pediatric practice (antibiotics and glucocorticoids) are also to blame for this situation. Low vitamin K status is much more frequent in newborns, due to both endogenous and exogenous insufficiencies. Just after birth vitamin K stores are low, and since human milk is relatively poor in this nutrient, breast-fed infants are at particular risk of a bleeding disorder called vitamin K deficiency bleeding. A pilot study showed that better vitamin K status is associated with lower rate of low-energy fracture incidence. An ongoing clinical trial is intended to address whether vitamin K2 and D3 supplementation might positively impact the biological process of bone healing. Vitamin K2 as menaquinone-7 (MK-7) has a documented history of safe and effective use. The lack of adverse effects of MK-7 makes it the ideal choice for supplementation by pregnant and nursing women and children, both healthy and suffering from various malabsorptions and health disorders, such as dyslipidemia, diabetes, thalassemia major (TM), cystic fibrosis (CF), inflammatory bowel diseases (IBD), and chronic liver diseases. Additionally, worthy of consideration is the use of vitamin K2 in obesity-related health outcomes.

Effect of Physiotherapy on the Promotion of Bone Mineralization in Preterm Infants. A Randomized Controlled Trial.

Preterm infants have a low level of bone mineralization compared to those born at term. The purpose of the present study was to investigate the effect of reflex locomotion therapy (RLT) on bone mineralization and growth in preterm infants and compare its effect against other physiotherapy procedures. 46 preterm infants born at 29-34 weeks were randomized to three groups: One group received RLT (n=17); other group received passive movements with gentle joint compression (n=14); and control group received massage (n=15). All treatments were carried out at the neonatal unit lasting one month. The main outcome measure was the bone mineralization measured with Tibial Speed of Sound (Tibial-SOS). All groups were similar in terms of gestational age (31.8±1.18), birth weight (1,583.41±311.9), and Tibia-SOS (1,604.7±27.9), at the beginning of the intervention. At the end of the study, significant differences were found among groups in Tibial-SOS [F(4,86)=2.77, p=0.049, ηp2 = 0.114] in benefit of RLT group. In conclusion, RLT has been effective in the improvement of Tibial-SOS, and has been more effective than other physical therapy modalities; therefore it could be considered as one of the most effective physiotherapeutic modalities for the prevention and treatment of osteopenia of prematurity.

Evaluation of Vitamin D-levels and Bone Mineral Density in Lymphoma and Solid Tumors of Childhood after Treatment

As the survival rate of childhood cancers increases, late effects of the treatment are becoming significantly more critical. The aim of this study is to evaluate serum 25 Hydroxy Vitamin D levels and bone mineral density (BMD) of patients with lymphoma and solid tumors. In this study, we included patients diagnosed with cancer in our pediatric oncology department between 2004 and 2013. These patients were all in remission and completed their treatment. Patient data were collected from corresponding medical files. Vitamin D levels, BMD measurements and other laboratory tests were performed prospectively. The study group consisted of 105 patients who completed treatment at least one year ago. Overall, 59.6 % of the subjects were found to have vitamin D levels less than 20 ng/mL. When the factors affecting vitamin D deficiency were evaluated, there were not any factor related to vitamin D deficiency. However, a positive correlation was observed in the blood sampling time, namely test results taken in summer months (April-September) were higher. In our study, bone mineralization disorder (low BMD) was found in 44.4% of the patients. In addition, we observed that bone health was significantly affected in children who recovered from cancer. 59.6 % of the vitamin D deficiency rate was found to be higher than the similar age group in our country. To our knowledge, this is also the lowest vitamin D levels in children with cancer compared to previously published studies. The rate of low BMD was determined as 44.4%, similar to earlier studies.

Suplementación de vitamina D en embarazadas y en mujeres con patologías ginecológicas

Apart from the well-known effects of vitamin D in maintaining calcium homeostasis and promoting bone mineralization, there is evidence to suggest that it modulates reproductive processes. About 70% adults have vitamin D deficiencies and there is a gap between recommended dietary intakes and deficient supply in general population. Observational studies show that vitamin D deficiency is a risk marker for reducing fertility and resulting adverse pregnancy. Evidence suggests that it could have beneficial effects on metabolic, hormonal and cellular parameters of polycystic ovarian syndrome, uterine myomatosis and endometriosis, while it seems to be associated with improvements in outcome of in vitro fertilization. The objective of this review was to evaluate the data of current literature on the role of vitamin D supplementation in pregnant women and in women with gynecological pathologies. Keywords: Vitamin D, Supplementation, Pregnancy, Endometriosis, Uterine myomatosis, Infertility.

Vitamin D in periodontology

Generalized periodontitis is one of the most common dental diseases that can lead to tooth mobility and tooth loss. Plaque is main etiological factor, but periodontitis is a multifactorial disease. There are data on the effect of vitamin D levels on generalized periodontitis. Vitamin D has a pleiotropic effect by affecting various links in the pathogenesis of generalized periodontitis. Stimulating the secretion of antimicrobial peptides, vitamin D increases local immunity and adversely affects periodontal pathogens; modulating the immune system, it affects inflammation in periodontal tissues and the immune response; increasing the absorption of calcium and reducing the release of parathyroid hormone, vitamin D stimulates bone mineralization. The high prevalence of vitamin D deficiency and insufficiency worldwide indicates the relevance of studying its effects on the human body in general and the oral cavity in particular.