The OPG/RANKL/RANK framework, along with its specific receptors, plays a crucial role in bone remodeling and the functioning of the central nervous system (CNS) and associated disorders. Recent research and investigations provide evidence that the components of osteoprotegerin (OPG), receptor activator of NF-kB ligand (RANKL), and receptor activator of NF-kB (RANK) are expressed in the CNS. The CNS structure encompasses cells involved in neuroinflammation, including local macrophages, inflammatory cells, and microglia that cross the blood-brain barrier. The OPG/RANKL/RANK trio modulates the neuroinflammatory response based on the molecular context. The levels of OPG/RANKL/RANK components can serve as biomarkers in the blood and cerebrospinal fluid. They act as neuroprotectants following brain injuries and also participate in the regulation of body weight, internal body temperature, brain ischemia, autoimmune encephalopathy, and energy metabolism. Although the OPG/RANKL/RANK system is primarily known for its role in bone remodeling, further exploring deeper into its multifunctional nature can uncover new functions and novel drug targets for diseases not previously associated with OPG/RANKL/RANK signaling.

The receptor-activator of NF-κB ligand (RANKL) and its specific receptor RANK have essential roles in regulating bone metabolism and the immune system. Besides, the RANKL/RANK system plays important roles in multiple physiological and pathophysiological processes such as mammary gland development during pregnancy, cancer development, and bone metastasis. While it has long been known that RANKL and RANK are expressed in the central nervous system (CNS), the physiological roles of RANKL/RANK system in the CNS and the underlying molecular mechanisms have been elucidated recently. Over the last decade, several reports showed that the central RANKL/RANK system plays important roles in regulating body temperature, brain ischemia, autoimmune encephalopathy, feeding behavior, and energy metabolism. In this review, it is provided an updated information regarding the roles of RANKL/RANK system in the CNS.

Almost a quarter century has passed since discovery of receptor activator of NF-κB ligand (RANKL). This discovery had a major impact on identification of mechanisms regulating osteoclast differentiation and function, establishment of a research field bridging bone and the immune system (osteoimmunology), and development of a fully human anti-RANKL neutralizing antibody (denosumab). Denosumab is now clinically available for treatment of osteoporosis and cancer-induced bone diseases in the US, Europe and many other countries, including Japan. Denosumab is a so-called blockbuster drug, with sales of 5.0 billion US dollars in 2019. This is a real success story from bench to bedside. In this review, the pivotal roles of the RANKL/RANK/OPG system in osteoclast differentiation and function are shown. RANKL is a ligand required for osteoclast generation, RANK is the receptor for RANKL, and osteoprotegerin (OPG) is a decoy receptor for RANKL. The review covers recent results showing the importance of RANKL on osteoblasts in regulation of osteogenesis and the role of RANKL-RANK dual signaling in coupling of bone resorption and formation, including demonstration of RANKL reverse signaling that we had previously hypothesized. Possible applications of anti-RANKL antibody in treatment of cancer are also discussed.

Obesity is a complex disorder that is influenced by genetic and environmental factors. DNA methylation is an epigenetic mechanism that is involved in development of obesity and its metabolic complications. The aim of this study was to investigate the association between the RANKL and c-Fos gene methylation on obesity with body mass index (BMI), lipid parameters, homeostasis model assessment of insulin resistance (HOMA-IR), plasma leptin, adiponectin and resistin levels. The study included 68 obese and 46 non-obese subjects. Anthropometric parameters, including body weight, body mass index, waist circumference, and waist-hip ratio, were assessed. Serum glucose, triglycerides (TG), total cholesterol, high-density lipoprotein cholesterol (HDL-C), and low-density lipoprotein cholesterol (LDL-C), plasma leptin, adiponectin and resistin levels were measured. Methylation status of RANKL and c-Fos gen were evaluated by MS-HRM. Statistically significant differences were observed between obese patients and the controls with respect to RANKL and c-Fos gene methylation status (p < 0.001). Also, statistically significant importance was observed RANKL gene methylation and increased level of leptin in obese subjects (p = 0.0081). At the same time, statistically significant association between methylation of c-Fos and increased level of adiponectin was observed in obese patients (p = 0.03) On the other hand, decreased level of resistin was observed where the c-Fos was unmetyladed in controls (p = 0.01). We conclude that methylation of RANKL and c-Fos genes have significant influences on obesity and adipokine levels. Based on literature this was the first study which shows the interactions between RANKL and c-Fos methylation and obesity.

To summarize the research progress on the calcitonin gene-related peptide (CGRP) and receptor activator of nuclear factor κB (RANK)/receptor activator of nuclear factor κB ligand (RANKL)/osteoprotegerin (OPG) system during bone reconstruction to provide theoretical basis for further research on the prevention and treatment of bone-related diseases.

The relevant research results at home and abroad in recent years were analyzed and summarized.

CGRP and RANK/RANKL/OPG system play important regulatory roles in the bone reconstruction.

At present, the research on the mechanism of CGRP and RANK/RANKL/OPG system in bone reconstruction is insufficient. Therefore, it is necessary to study further on the process and interrelation of CGRP and RANK/RANKL/OPG system in bone reconstruction to confirm their mechanism, which will bring new ideas and methods for the treatment of bone related diseases in clinic.

Serum calcium (Ca) is maintained in a narrow range through regulation of Ca metabolism in the intestine, kidney, and bone. Calcium is incorporated and resorbed from bone during bone remodeling via cellular processes as well as by exchange. Both routes contribute to calcium homeostasis. To assess the magnitude of bone turnover contribution to calcium homeostasis we labeled bone with a Ca tracer and measured Ca release following stimulation or suppression of bone resorption. Young growing male rats (n = 162) were dosed with ⁴⁵Ca to label skeletal Ca. After a one-month period to allow the label to incorporate into the skeleton, rats were treated with a bone resorption antagonist (OPG), a bone resorption agonist (RANKL), or vehicle control (PBS). Serum and urine ⁴⁵Ca and total Ca, and serum TRACP5b (a bone resorption biomarker), were monitored for 45 days following treatment. Tracer data were analyzed by a compartmental model using WinSAAM to quantify dynamic changes in Ca metabolism and identify sites of change following treatment. In RANKL treated rats, both serum ⁴⁵Ca and serum TRACP5b were increased by >70% due to a 25-fold increase in bone resorption. In OPG treated rats, both serum ⁴⁵Ca and serum TRACP5b were suppressed by >70% due to a 75% decrease in bone resorption, a 3-fold increase in bone formation, and a 50% increase in absorption. Because TRACP5b and ⁴⁵Ca responded similarly, we conclude that Ca release from bone into serum occurs mostly via osteoclast-mediated bone resorption. However, because serum Ca concentration did not change with altered resorption in response to either RANKL or OPG treatment, we also conclude that serum Ca concentration under normal dietary conditions in young growing male rats is maintained by processes in addition to cellular bone resorption.

The receptor activator of nuclear factor-κB ligand (RANKL) modulates energy metabolism. However, how RANKL regulates energy homeostasis is still not clear. This study aims to investigate the central mechanisms by which central administration of RANKL inhibits food intake and causes weight loss in mice. We carried out a systematic and in-depth analysis of the neuronal pathways by which RANKL mediates catabolic effects. After intracerebroventricle (i.c.v.) injection of RANKL, the expression of neuropeptide Y (NPY) mRNA in the Arc was significantly decreased, while the CART mRNA expression dramatically increased in the Arc and DMH. However, the agouti-related protein (AgRP) and pro-opiomelanocortin (POMC) mRNA had no significant changes compared with control groups. Together, the results suggest that central administration of RANKL reduces food intake and causes weight loss via modulating the hypothalamic NPY/CART pathways.

This study investigated the effects of rifampicin-modulated P-glycoprotein (P-gp) and cytochrome P450 (CYP450) activity on the development of steroid-induced osteonecrosis of the femoral head. Thirty-two rabbits were equally divided into four groups: control group, oral administration group, intramuscular injection group, and local release group, in which rifampicin-loaded artificial bone graft was implanted in the left femur cavity and blank bone graft was implanted in the right femur cavity. Dexamethasone was given 1 week after rifampicin administration. Peripheral P-gp activity and hepatic CYP450 content were investigated 4 weeks later. Hematoxylin and eosin, Massson, and tetracycline-fluorescence staining of the femoral head were compared. In vitro, the effects of intracellular dexamethasone concentration modulated by P-gp on osteoprotegerin (OPG)/receptor activator of nuclear factor κB ligand (RANKL) expression and differentiation of mesenchymal stem cells were further investigated. Peripheral P-gp activity and hepatic CYP450 content in the oral administration group and the intramuscular injection group were significantly higher than those in the local release group. P-gp activity of mesenchymal stem cells in rifampicin-implanted femoral head was significantly higher than that in the blank control. Histological study showed that rifampicin could prevent steroid-induced bone loss and lipid formation, and promote new bone formation and maturation. In vitro study confirmed that intracellular dexamethasone concentration modulated by P-gp could influence the OPG/RANKL ratio and the differentiation of mesenchymal stem cells. Enhanced levels of peripheral P-gp and hepatic CYP450 can reduce the incidence of steroid-induced osteonecrosis of the femoral head. P-gp activity locally enhanced by rifampicin decreases the intracellular steroid concentration, but rifampicin does not have significant effects on peripheral P-gp and hepatic CYP450.

Osteoporosis is a systemic skeletal disease with the high incidence, serious complications, financial burden, and heavily decrease in living quality.

Proliferation of osteoblast was tested by 2,3-bis-(2-methoxy-4-nitro-5-sulfophenyl)-2H-tetrazolium-5-carboxanilide (XTT) method, alkaline phosphatase (ALP) activity of osteoblasts was tested by ALP REAGENT, Calcium level was determined by a colorimetric assay, mRNA expression of phosphoinositide-3 kinase (PI3K), 3-phosphoinositide-dependent protein kinase 1 (PDK1), Akt, Caspase-3, Caspase-7, Caspase-9, osteocalcin (OCN), Osterix and Runx2 of osteoblasts was tested by RNA preparation and quantitative reverse transcription polymerase chain reaction (RT-PCR), and protein expression of phospho-PI3K, phospho-PDK1 and phospho-Akt was measured by Western Blot analysis.

In osteoporosis model rats, it found that mRNA expression of PI3K, PDK1 and Akt showed no changes while protein expression of phospho-PI3K, phospho-PDK1 and phospho-Akt in bone tissue was decreased dramatically. To further characterize the molecular mechanisms that regulate osteoporosis, we examined the contribution of the PI3K/Akt cell signaling pathway in cultured osteoblasts. It suggested that, the blockade of PI3K activation by LY294002, a specific inhibitor of the PI3K/Akt signaling pathway in osteoblasts, heavily inhibited cell proliferation, ALP activity, calcium accumulation, and mRNA expression of OCN, Osterix and Runx2. However, mRNA expression of Caspase-3 and Caspase-9 was promoted accordingly.

The in vivo and in vitro studies indicated that the PI3K/Akt cell signaling pathway is involved in the inhibition of osteoporosis through promoting osteoblast proliferation, differentiation and bone formation.

We previously demonstrated that dietary vitamin D(3) at 10x the adequate intake (AI) attenuates the decline in functional capacity in the G93A mouse model of ALS. We hypothesized that higher doses would elicit more robust changes in functional and disease outcomes.

To determine the effects of dietary vitamin D(3) at 50xAI on functional outcomes (motor performance, paw grip endurance) and disease severity (clinical score), as well as disease onset, disease progression and lifespan in the transgenic G93A mouse model of ALS.

Starting at age 25 d, 100 G93A mice (55 M, 45 F) were provided ad libitum with either an adequate (AI; 1 IU D(3)/g feed) or high (HiD; 50 IU D(3)/g feed) vitamin D(3) diet.

HiD females consumed 9% less food corrected for body weight vs. AI females (P = 0.010). HiD mice had a 12% greater paw grip endurance over time between age 60-141 d (P = 0.015), and a 37% greater score during disease progression (P = 0.042) vs. AI mice. Although HiD females had a non-significant 31% greater CS prior to disease onset vs. AI females, they exhibited a significant 20% greater paw grip endurance AUC (P = 0.020) when corrected for clinical score.

Dietary D(3) supplementation at 50x the adequate intake attenuated the decline in paw grip endurance, but did not influence age at disease onset, hindlimb paralysis or endpoint in the transgenic G93A mouse model of ALS. Furthermore, females may have reached the threshold for vitamin D(3) toxicity as evidence by reduced food intake and greater disease severity prior to disease onset.