Suppression of bone resorption by Mori Radicis Cortex through NFATc1 and c-Fos signaling-mediated inhibition of osteoclast differentiation.

BACKGROUND: Mori Radicis Cortex (MRC) is the root bark of the mulberry family as Morus alba L. In Korea, it is known as "Sangbaegpi". Although MRC has demonstrated anti-inflammatory and antioxidant effects, its specific mechanisms of action and impact on osteoporosis remain poorly understood. METHODS: To investigate the antiosteoporosis effect of MRC, we examined the level of osteoclast differentiation inhibition in receptor activator of nuclear factor kappa-Β ligand (RANKL)-induced-RAW 264.7 cells and animal models of ovariectomy (OVX) with MRC. Serum analysis in OVX animals was investigated by enzyme-linked immunosorbent assay (ELISA), and bone density analysis was confirmed by micro-computed tomography (micro-CT). The expression analysis of nuclear factor of activated T cells 1 (NFATc1) was confirmed by immunohistochemistry (IHC) in femur tissue. In addition, osteoclast differentiation inhibition was measured using tartrate-resistant acid phosphatase (TRAP). mRNA analysis was performed using reverse transcription-polymerase chain reaction (RT-PCR), and the protein expression analysis was investigated by western blot. RESULTS: Micro-CT analysis showed that MRC effectively inhibited bone loss in the OVX-induced rat model. MRC also inhibited the expression of alkaline phosphatase (ALP) and TRAP in serum. Histological analysis showed that MRC treatment increased bone density and IHC analysis showed that MRC significantly inhibited the expression of NFATc1. In RANKL-induced-RAW 264.7 cells, MRC significantly reduced TRAP activity and actin ring formation. In addition, MRC significantly inhibited the expression of NFATc1 and c-Fos, and suppressed the mRNA expression. CONCLUSION: Based on micro-CT, serum and histological analysis, MRC effectively inhibited bone loss in an OVX-induced rat model. In addition, MRC treatment suppressed the expression of osteoclast differentiation, fusion, and bone resorption markers through inhibition of NFATc1/c-Fos expression in RANKL-induced RAW 264.7 cells, ultimately resulting in a decrease in osteoclast activity. These results demonstrate that MRC is effective in preventing bone loss through inhibiting osteoclast differentiation and activity.

Exploring the Anti-Osteoporotic Potential of Daucosterol: Impact on Osteoclast and Osteoblast Activities.

Osteoporosis is a debilitating condition characterized by reduced bone mass and density, leading to compromised structural integrity of the bones. While conventional treatments, such as bisphosphonates and selective estrogen receptor modulators (SERMs), have been employed to mitigate bone loss, their effectiveness is often compromised by a spectrum of adverse side effects, ranging from gastrointestinal discomfort and musculoskeletal pain to more severe concerns like atypical fractures and hormonal imbalances. Daucosterol (DC), a natural compound derived from various plant sources, has recently garnered considerable attention in the field of pharmacology. In this study, we investigated the anti-osteoporosis potential of DC by characterizing its role in osteoclasts, osteoblasts, and lipopolysaccharide (LPS)-induced osteoporosis. The inhibitory effect of DC on osteoclast differentiation was determined by tartrate-resistant acid phosphatase (TRAP) staining, F-actin ring formation by fluorescent staining, and bone resorption by pit formation assay. In addition, the calcification nodule deposition effect of osteoblasts was determined by Alizarin red S staining. The effective mechanisms of both cells were verified by Western blot and reverse transcription polymerase chain reaction (RT-PCR). To confirm the effect of DC in vivo, DC was administered to a model of osteoporosis by intraperitoneal administration of LPS. The anti-osteoporosis effect was then characterized by micro-CT and serum analysis. The results showed that DC effectively inhibited osteoclast differentiation at an early stage, promoted osteoblast activity, and inhibited LPS-induced bone density loss. The results of this study suggest that DC can treat osteoporosis through osteoclast and osteoblast regulation, and therefore may be considered as a new therapeutic alternative for osteoporosis patients in the future.

Dolichos Lablab Linné Inhibits Bone Density Loss and Promotes Bone Union in Senile Osteoporosis through Osteogenesis.

As populations continue to age, osteoporosis has emerged as an increasingly critical concern. Most advancements in osteoporosis treatment are predominantly directed toward addressing abnormal osteoclast activity associated with menopause, with limited progress in developing therapies that enhance osteoblast activity, particularly in the context of aging and fractures, and serious side effects associated with existing treatments have highlighted the necessity for natural-product-based treatments targeting senile osteoporosis and fractures. Dolichos lablab Linné (DL) is a natural product traditionally used for gastrointestinal disorders, and its potential role in addressing bone diseases has not been extensively studied. In this research, we investigated the anti-osteoporosis and bone-union-stimulating effects of DL using the SAMP6 model, a naturally aged mouse model. Additionally, we employed MC3T3-E1 cells to validate DL's osteoblast-promoting effect and to assess the involvement of core mechanisms such as the BMP-2/Smad and Wnt/ß-catenin pathways. The experimental results revealed that DL promoted the formation of osteoblasts and calcified nodules by upregulating both the BMP-2/Smad and Wnt/ß-catenin mechanisms. Based on its observed effects, DL demonstrated the potential to enhance bone mineral density in aged osteoporotic mice and promote bone union in fractured mice. These findings indicate the promising therapeutic potential of DL for the treatment of osteoporosis and bone-related conditions, thus warranting further investigation and potential clinical applications.

Persicae Semen Promotes Bone Union in Rat Fractures by Stimulating Osteoblastogenesis through BMP-2 and Wnt Signaling.

Fractures cause extreme pain to patients and impair movement, thereby significantly reducing their quality of life. However, in fracture patients, movement of the fracture site is restricted through application of a cast, and they are reliant on conservative treatment through calcium intake. Persicae semen (PS) is the dried mature seeds of Prunus persica (L.) Batsch, and in this study the effects of PS on osteoblast differentiation and bone union promotion were investigated. The osteoblast-differentiation-promoting effect of PS was investigated through alizarin red S and Von Kossa staining, and the regulatory role of PS on BMP-2 (Bmp2) and Wnt (Wnt10b) signaling, representing a key mechanism, was demonstrated at the protein and mRNA levels. In addition, the bone-union-promoting effect of PS was investigated in rats with fractured femurs. The results of the cell experiments showed that PS promotes mineralization and upregulates RUNX2 through BMP-2 and Wnt signaling. PS induced the expression of various osteoblast genes, including Alpl, Bglap, and Ibsp. The results of animal experiments show that the PS group had improved bone union and upregulated expression of osteogenic genes. Overall, the results of this study suggest that PS can promote fracture recovery by upregulating osteoblast differentiation and bone formation, and thus can be considered a new therapeutic alternative for fracture patients.

Exposure to serum perfluoroalkyl substances and biomarkers of liver function: The Korean national environmental health survey 2015-2017.

BACKGROUND: Exposure to perfluoroalkyl substances (PFAS) may increase the risk of liver disease by disrupting cholesterol and lipid synthesis/metabolism, leading to higher liver-enzyme concentrations. However, most studies assessing association between PFAS and liver enzymes focused on individual PFAS. Moreover, PFAS concentrations differ based on sex and obesity status, and it remains unclear whether these factors affect associations with liver function. Therefore, we examined the association between exposure to both individual and combined PFAS and liver-function biomarkers and assessed sex and obesity as effect modifiers in Korean adults. METHODS: We measured serum concentrations of the five most abundant PFAS (PFOA, PFOS, PFHxS, PFDA, PFNA) and three liver enzymes (alanine transaminase [ALT], aspartate aminotransferase [AST], γ-glutamyl transferase [GGT]) in 1404 adults from the Korean National Environmental Health Survey Cycle 3, 2015-2017. We used linear regression to evaluate associations between individual PFAS and liver-function biomarkers, assessing sex and obesity as possible effect modifiers, and performed Bayesian kernel machine regression and quantile g-computation to evaluate the overall effect of PFAS mixture on biomarkers of liver function. RESULTS: Among 1404 Korean adults, all five PFAS were detected. Geometric mean concentration was highest for PFOS (16.11 µg/L), followed by PFOA (5.83 µg/L), PFHxS (2.21 µg/L), PFNA (2.03 µg/L), and PFDA (1.06 µg/L). In multivariable linear regression, all PFAS were positively associated with ALT, AST, and GGT; 2-fold increase in each PFAS was associated with 3.4-8.6% higher ALT, 2.4-4.6% higher AST, and 4.6-11.1% higher GGT (all p < 0.05). Positive associations for PFOA, PFDA, and PFNA with AST were stronger in men, and positive associations for PFOS with ALT and GGT were stronger in women. Compared to obese participants, nonobese participants had higher average percent changes in each enzyme, particularly GGT, when individual PFAS concentration doubled. Additionally, increased exposure to PFAS mixtures was associated with higher ALT, AST, and GGT. In quantile g-computations, simultaneous quartile increase in all PFAS was significantly associated with 6.9% (95%CI: 3.7, 10.2) higher ALT, 4.5% (95%CI: 2.4, 6.6) higher AST, and 8.3% (95%CI: 3.7, 13.1) higher GGT levels, on average. CONCLUSIONS: Exposure to individual and combined PFAS is associated with higher liver enzymes in Korean adults, providing additional evidence for the association between PFAS exposure and risk of liver disease.

A Study on Safety Management Plan for Recycling of Medium-Contact Wastes via Ecotoxicity Assessment.

In South Korea, hazardous characteristics of wastes to be recycled are managed through the "Environmental Impact Assessment of Recycling" system. The ecotoxicity of medium-contact recyclable wastes, that is, those in contact with soil, groundwater, surface water, etc., is managed according to this system and is determined based on whether or not they exceed an ecotoxicity value (TU) of 2.0. The ecotoxicity of wastes is tested and determined by using pretreated eluate samples according to the Official Wastes Test Standard and applying the Official Water Pollution Process Test Standard. However, no ecotoxicity management limits are stipulated for medium-contact recycling using wastes in numerous other countries. This study aims to evaluate applicability and safety of the ecotoxicity test for wastes used in medium-contact recycling and establish an efficient management plan for hazardous characteristic wastes. Target wastes for the survey were selected based on the Wastes Control Act in South Korea. Nine types of waste were selected, which are representative types of wastes to which ecotoxicity is applied. In order to secure the representativeness of the target samples, a total of 45 samples were collected by selecting 5 cases each of the 9 waste types in consideration of the type of industry and amount of waste generated. Limit exceedance was calculated for each category of hazardous substances (leaching, total content), pH, and ecotoxicity of a total of 45 samples, and was found to increase in the order of leaching 2.22% < pH 9.09% < content 31.11% < ecotoxicity 37.21%. This indicates that the limit exceedance was maximum in the ecotoxicity category. Therefore, the application of ecotoxicity limit is efficient for identifying and comprehensively managing the environmental impacts of various types of hazardous substances contained in wastes from the perspective of comprehensive toxicity.

Dendrobium nobile Lindley Administration Attenuates Atopic Dermatitis-like Lesions by Modulating Immune Cells.

Atopic dermatitis (AD) is a chronic inflammatory skin disease that can significantly affect daily life by causing sleep disturbance due to extreme itching. In addition, if the symptoms of AD are severe, it can cause mental disorders such as ADHD and suicidal ideation. Corticosteroid preparations used for general treatment have good effects, but their use is limited due to side effects. Therefore, it is essential to minimize the side effects and study effective treatment methods. Dendrobium nobile Lindley (DNL) has been widely used for various diseases, but to the best of our knowledge, its effect on AD has not yet been proven. In this study, the inhibitory effect of DNL on AD was confirmed in a DNCB-induced Balb/c mouse. In addition, the inhibitory efficacy of inflammatory cytokines in TNF-α/IFN-γ-induced HaCaT cells and PMACI-induced HMC-1 cells was confirmed. The results demonstrated that DNL decreased IgE, IL-6, IL-4, scratching behavior, SCORAD index, infiltration of mast cells and eosinophils and decreased the thickness of the skin. Additionally, DNL inhibited the expression of cytokines and inhibited the MAPK and NF-κB signaling pathways. This suggests that DNL inhibits cytokine expression, protein signaling pathway, and immune cells, thereby improving AD symptoms in mice.

Profile of Environmental Chemicals in the Korean Population-Results of the Korean National Environmental Health Survey (KoNEHS) Cycle 3, 2015-2017.

The Korean National Environmental Health Survey (KoNEHS) program provides useful information on chemical exposure, serves as the basis for environmental health policies, and suggests appropriate measures to protect public health. Initiated on a three-year cycle in 2009, it reports the concentrations of major environmental chemicals among the representative Korean population. KoNEHS Cycle 3 introduced children and adolescents into the analysis, where the blood and urine samples of 6167 participants were measured for major metals, phthalates, phenolics, and other organic compounds. Lead, mercury, cadmium, metabolites of DEHP and DnBP, and 3-phenoxybenzoic acid levels of the Korean adult population tended to decrease compared to previous survey cycles but remained higher than those observed in the US or Canada. Both bisphenol A (BPA) and trans,trans-muconic acid concentrations have increased over time. Heavy metal concentrations (blood lead, and cadmium) in children and adolescents were approximately half that of adults, while some organic substances (e.g., phthalates and BPA) were high. BPA showed higher levels than in the US or Canada, whereas BPF and BPS showed lower detection rates in this cycle; however, as these are increasingly used as a substitute for BPA, further research is necessary. As environmental chemicals may affect childhood health and development, additional analyses should assess exposure sources and routes through continuous observations.

Lead, mercury, and cadmium exposures are associated with obesity but not with diabetes mellitus: Korean National Environmental Health Survey (KoNEHS) 2015-2017.

BACKGROUND: Associations of heavy metal exposures with obesity and obesity-related traits have been suggested, while those with nonalcoholic fatty liver disease (NAFLD) and diabetes mellitus (DM) are often inconsistent. METHODS: This study included 3787 adults aged ≥19 years who participated in the Korean National Environmental Health Survey 2015-2017, and investigated the association of toxic heavy metals with metabolic diseases. Lead (Pb), mercury (Hg), and cadmium (Cd) were measured either in urine (uHg, uCd) or total blood (bPb, bHg). Body mass index (BMI) was calculated, and DM cases were identified through a self-answered medication history. Hepatic Steatosis Index (HSI) as a surrogating index of NAFLD, was calculated using hepatic enzyme measurements, including aspartate aminotransferase (AST) and alanine aminotransferase (ALT). RESULTS: Adults in the highest quartile of bPb, bHg, and uHg showed significantly elevated odds of obesity (BMI ≥25 kg/m2), compared to the lowest quartile (OR 1.58 for bPb, 1.92 for bHg, and 1.81 for uHg). HSI was positively correlated with bHg, uHg, and uCd concentrations. The odds of NAFLD (HSI ≥36) were also increased with increasing quartile of bHg, uHg, and uCd concentrations. For DM, bPb showed a significant negative association, while bHg and uCd exhibited non-monotonic and inconclusive associations. CONCLUSIONS: Among the general adult population of Korea, both Pb and Hg exposures were associated with an increased risk of obesity. In addition, both Hg and Cd exposures were associated with increased odds of NAFLD. These metals, however, were not associated with an increased risk of DM.

Exposure to polycyclic aromatic hydrocarbons and volatile organic compounds is associated with a risk of obesity and diabetes mellitus among Korean adults: Korean National Environmental Health Survey (KoNEHS) 2015-2017.

Environmental pollutants have been known to increase the risks of not only respiratory and cardiovascular disease but also metabolic diseases such as obesity and diabetes mellitus (DM). Polycyclic aromatic hydrocarbons (PAHs) and volatile organic compounds (VOCs) such as benzene and toluene are major constituents of environmental pollution. In the present study, we employed the population of the Korean National Environmental Health Survey (KoNEHS) Cycle 3 conducted between 2015 and 2017, and assessed the associations of urinary biomarkers for PAHs and VOCs exposure with obesity and DM. A total of 3787 adult participants were included and the urinary concentrations of four PAH metabolites and two VOC metabolites were measured. For correcting urine dilution, a covariate-adjusted standardization method was used. The highest quartiles of urinary 2-hydroxynaphthalene (2-NAP) [OR (95% confidence interval (CI)) = 1.46 (1.13, 1.87)] and sum of PAH metabolites [OR (95% CI) = 1.45 (1.13, 1.87)] concentrations were associated with a higher risk of obesity [body mass index (BMI)≥25 kg/m2]. BMI was positively associated with urinary 2-NAP [ß (95% CI) = 0.25 (0.09, 0.41), p = 0.003] and sum of PAH metabolites [ß (95% CI) = 0.29 (0.08, 0.49), p = 0.006] concentrations. The risk of DM was increased with increasing quartile of 2-hydroxyfluorene (2-OHFlu) and trans, trans-muconic acid (t,t-MA) (p for trend<0.05 and < 0.001, respectively). The highest quartile of t,t-MA showed a significantly higher risk of DM [OR (95% CI) = 2.77 (1.74, 4.42)] and obesity [OR (95% CI) = 1.42 (1.06, 1.90)]. Urinary t,t,-MA level was positively associated with BMI [(ß (95% CI) = 0.51 (0.31, 0.71), p < 0.001] and non-alcoholic fatty liver disease index [(ß (95% CI) = 0.09 (0.06, 0.12), p < 0.001]. In conclusion, the benzene metabolites t,t-MA and PAH metabolite 2-OHFlu were associated with an increased risk of DM. Urinary biomarkers for PAHs and VOCs were positively associated with BMI in the Korean adult population. Further studies to validate these observations in other populations are warranted.

Lycopus lucidus Turcz ameliorates DNCB­induced atopic dermatitis in BALB/c mice.

Atopic dermatitis (AD) is a chronic inflammatory allergic skin disease, characterized by pruritic and eczematous skin lesions. Lycopus lucidus Turcz (LLT) is a perennial herb that has been reported to have various biological properties, including effects on blood circulation, as well as anti­inflammatory, antioxidant, anti­vascular inflammation and wound­healing effects. However, whether LLT improves dermatitis and the underlying mechanisms has yet to be determined. The aim of the present study was to determine whether LLT can improve 2,4­dinitrochlorobenzene (DNCB)­induced dermatitis and to verify the inhibitory effect of LLT on the expression of chemokines and pro­inflammatory cytokines in the HaCaT immortalized keratinocyte cell line. In addition, the anti­inflammatory function of LLT in RAW264.7 mouse macrophages was investigated. In the DNCB­induced AD mouse model, LLT inhibited infiltration by mast cells, eosinophils and CD8+ cells in the dorsal skin tissue of AD mice, and suppressed the expression of IgE and IL­6 in serum. In addition, LLT inhibited the phosphorylation of ERK and JNK, as well as NF­κB in skin tissue. In the HaCaT cell model induced by TNF­α/IFN­Î³, LLT inhibited the expression of thymus and activation­regulated chemokine, granulocyte­macrophage colony­stimulating factor, monocyte chemoattractant protein­1, TNF­α and IL­1ß, whilst inhibiting the phosphorylation of NF­κB. In addition, in the lipopolysaccharide­induced RAW 264.7 cell inflammation model, LLT inhibited the expression of TNF­α and IFN­Î³, the nuclear translocation of NF­κB and the phosphorylation of ERK and JNK. These results suggested that LLT may be a promising candidate for the treatment of inflammatory dermatitis.

Albiflorin Promotes Osteoblast Differentiation and Healing of Rat Femoral Fractures Through Enhancing BMP-2/Smad and Wnt/ß-Catenin Signaling.

Fracture healing is related to osteogenic differentiation and mineralization. Recently, due to the unwanted side effects and clinical limitations of existing treatments, various natural product-based chemical studies have been actively conducted. Albiflorin is a major ingredient in Paeonia lactiflora, and this study investigated its ability to promote osteogenic differentiation and fracture healing. To demonstrate the effects of albiflorin on osteoblast differentiation and calcified nodules, alizarin red S staining and von Kossa staining were used in MC3T3-E1 cells. In addition, BMP-2/Smad and Wnt/ß-catenin mechanisms known as osteoblast differentiation mechanisms were analyzed through RT-PCR and western blot. To investigate the effects of albiflorin on fracture healing, fractures were induced using a chainsaw in the femur of Sprague Dawley rats, and then albiflorin was intraperitoneally administered. After 1, 2, and 3 weeks, bone microstructure was analyzed using micro-CT. In addition, histological analysis was performed by staining the fractured tissue, and the expression of osteogenic markers in serum was measured. The results demonstrated that albiflorin promoted osteoblastogenesis and the expression of RUNX2 by activating BMP-2/Smad and Wnt/ß-catenin signaling in MC3T3-E1 cells. In addition, albiflorin upregulated the expression of various osteogenic genes, such as alkaline phosphatase, OCN, bone sialoprotein, OPN, and OSN. In the femur fracture model, micro-CT analysis showed that albiflorin played a positive role in the formation of callus in the early stage of fracture recovery, and histological examination proved to induce the expression of osteogenic genes in femur tissue. In addition, the expression of bone-related genes in serum was also increased. This suggests that albiflorin promotes osteogenesis, bone calcification and bone formation, thereby promoting the healing of fractures in rats.

Effects of chloroform fraction of Fritillariae Thunbergii Bulbus on atopic symptoms in a DNCB-induced atopic dermatitis-like skin lesion model and in vitro models.

ETHNOPHARMACOLOGICAL RELEVANCE: Fritillariae thunbergii Bulbus (FT), knowns as "Jeolpaemo ()" in Korean traditional medicine, is a perennial plant belonging to the Liliaceae family and has been used to treat symptoms such as cough, sputum formation, and purulent pneumonia. Owing to its effects of lowering heat, removing sputum, and reducing swelling, the plant has also been used as an external prescription medicine to treat inflammation. AIM OF THE STUDY: To analyze the anti-inflammatory effects of FT-ethanol extract (FT-Et) and FT-chloroform fraction extract (FT-Cl) on 1-chloro-2,4-dinitrobenzene (DNCB)-induced atopic dermatitis (AD) in vivo and in vitro. MATERIALS AND METHODS: The effect of FT-Et and FT-Cl on AD was observed using an AD-like skin lesion model induced by DNCB in vivo. HaCaT and RBL2H3 cells were used to determine the effects of FT-Et and FT-Cl in vitro. After inducing AD-like skin lesions in vivo, FT was topically applied to the skin lesion for 35 days. Epidermal thickness, dermal thickness, scratching behavior, infiltration of inflammatory cells, and expression of skin barrier proteins were measured. TARC, MDC, and IL-4 levels were analyzed using ELISA in HaCaT cells. Beta-hexosaminidase and IL-4 levels were measured in RBL2H3 cells. The expression of filaggrin (FLG), loricrin (LOR), involucrin (INV), and aquaporin-3(AQP-3) was measured by PCR. Phosphorylation of MAPKs was analyzed using Western blot technique. RESULTS: FT-Cl significantly reduced ear swelling, scratching behavior, SCORAD index, epidermal thickness, infiltration of inflammatory cells, and loss of skin barrier proteins. FT-Et inhibited the infiltration of mast cells and CD8+ cells and decreased the loss of skin barrier proteins. In TNF-α/IFN-γ-stimulated HaCaT cells, FT-Cl inhibited TRAC, MDC, and IL-4 expression and upregulated the expression of FLG, INV, and AQP-3, whereas FT-Et inhibited the expression of TRAC and MDC and increased the expression of FLG, INV, and AQP-3 at high concentrations. In RBL2H3, FT-Cl downregulated ß-hexosaminidase and IL-4 expression. In addition, FT-Cl inhibited the phosphorylation of ERK and p-38 in HaCaT and RBL2H3 cells. CONCLUSIONS: Collectively, FT-Cl showed better effect than FT-Et in vivo and in vitro. These results suggest that a specific component present in FT-Cl acted against AD. Future research should focus on the analysis of components contained in FT-Cl and the anti-inflammatory effects of the active ingredient.

Lycii radicis cortex inhibits glucocorticoid­induced bone loss by downregulating Runx2 and BMP­2 expression.

Lycii radicis cortex (LRC) has been used to regulate high blood pressure, body temperature, pain and bone disorders in East Asia. Glucocorticoids (GCs), also known as steroids, are potent immunity regulators widely used in the treatment of inflammatory diseases. However, despite their effectiveness, GC usage is strictly controlled due to severe side­effects, such as osteoporosis. However, further research is required as to date, at least to the best of our knowledge, there is no appropriate model to overcome secondary osteoporosis as a side­effect of GC use. Thus, the aim of the present study was to establish an experimental model of osteoporosis induced by GC. Furthermore, the present study aimed to establish the research methodology for medical evaluations of the effectiveness and side­effects of GCs. A secondary osteoporosis animal model was established, and the animals were divided into two groups as follows: The allergic contact dermatitis (ACD)­induced group and the non­ACD­induced group. In the ACD­induced group, a GC topical application group was compared with a GC subcutaneous injection group. The results revealed that the presence of ACD affected the induction of GC­mediated osteoporosis. Therefore, the group exhibiting induced ACD that was treated with a topical application of GC was selected for examining the side­effects of GCs. The effects of LRC on secondary osteoporosis were confirmed in vivo and in vitro. The results indicated that LRC regulated dexamethasone­induced osteoblast apoptotic markers, including caspase­6, caspase­9, X­linked inhibitor of apoptosis, apoptosis inhibitor 1 and apoptosis inhibitor 2, and increased the expression of osteoblast differentiation­related genes, such as Runt­related transcription factor 2 and bone morphogenetic protein 2 in the MC3T3E­1 cell line. LRC also significantly reduced GC­induced osteoporosis and exerted anti­inflammatory effects in vivo. In addition, LRC inhibited the reduction of calbindin­D28k in the kidney. Overall, the results of the present study suggest that the use of LRC alleviates GC­induced secondary osteoporosis.

Effects of Melandrium firmum Rohrbach on RANKL­induced osteoclast differentiation and OVX rats.

Osteoporosis is a systemic skeletal disease characterized by reduced bone mineral density (BMD), which results in an increased risk of fracture. Melandrium firmum (Siebold & Zucc.) Rohrbach (MFR), 'Wangbulryuhaeng' in Korean, is the dried aerial portion of Melandrii Herba Rohrbach, which is a member of the Caryophyllaceae family and has been used to treat several gynecological conditions as a traditional medicine. However, to the best of our knowledge, the effect of MFR on osteoclast differentiation and osteoporosis has not been assessed. To evaluate the effects of MFR on osteoclast differentiation, tartrate­resistant acid phosphatase staining, actin ring formation and bone resorption assays were used. Additionally, receptor activator of nuclear factor­κB ligand­induced expression of nuclear factor of activated T cell, cytoplasmic 1 (NFATc1) and c­Fos were measured using western blotting and reverse transcription­PCR. The expression levels of osteoclast­related genes were also examined. To further investigate the anti­osteoporotic effects of MFR in vivo, an ovariectomized (OVX) rat model of menopausal osteoporosis was established. Subsequently, the femoral head was scanned using micro­computed tomography. The results revealed that MFR suppressed osteoclast differentiation, formation and function. Specifically, MFR reduced the expression levels of osteoclast­related genes by downregulating transcription factors, such as NFATc1 and c­Fos. Consistent with the in vitro results, administration of MFR water extract to OVX rats reduced BMD loss, and reduced the expression levels of NFATc1 and cathepsin K in the femoral head. In conclusion, MFR may contribute to alleviate osteoporosis­like symptoms. These results suggested that MFR may exhibit potential for the prevention and treatment of postmenopausal osteoporosis.

Urinary parabens and their potential sources of exposure among Korean children and adolescents: Korean National Environmental Health Survey 2015-2017.

Parabens are used as a preservative in several consumer products including cosmetics, personal care products, and medicinal products. These chemicals have been suspected for estrogenicity and potential adverse endocrine outcomes in humans. For the first time, exposure profiles and potential sources of major parabens are investigated for a nationally representative population of children and adolescents of Korea. In addition, major determinants of urinary paraben levels were identified. For this purpose, the children, and adolescents (n = 2355, 3-18 years of age) who participated in the Korean National Environmental Health Survey cycle 3 (2015-2017) were studied. Adjusted multiple linear regression models were employed to investigate the relationships of several potential demographic and behavioral determinants of exposure, with the urinary levels of three parabens; methyl, ethyl, and propyl paraben. Methyl and propyl paraben levels of the Korean children and adolescents were comparable to those of the US, but the high exposure group (95th percentile) showed much higher levels of exposure. Moreover, urinary ethyl paraben levels are always higher than those of other countries. The uses of personal care products including liquid soaps, fragrance products, nail polish, or antiseptic products were significantly associated with urinary paraben levels. In addition, dietary sources such as fast food and canned food consumption were identified as major contributors to ethyl paraben levels. For methyl and propyl parabens, the use of fever medications and ointments were identified as major determinants of the exposure, especially among the younger children of 3-5 years of age. These observations are related to the Korean regulations that permit the use of the parabens as preservatives in foods and medications. The findings demonstrate that the exposure profile of parabens among Korean children are unique, and mitigation efforts for some parabens are required in Korea. Further studies are warranted to confirm the exposure sources of parabens and to develop mitigation measures among Korean children and adolescents.

First nationwide exposure profile of major persistent organic pollutants among Korean adults and their determinants: Korean National Environmental Health Survey Cycle 3 (2015-2017).

Since 2009, Korea has measured the exposure levels of major environmental chemicals and heavy metals among representative adult populations through the Korean National Environmental Health Survey (KoNEHS). However, exposure to persistent organic pollutants (POPs) has never been assessed. This study reports the serum concentrations of twenty-four POPs and their influencing factors for Korean adults (n = 1295) who participated in the KoNEHS Cycle 3 (2015-2017). The POPs included seven organochlorine pesticides (OCPs), eleven polychlorinated biphenyls (PCBs), and six polybrominated diphenyl ethers (PBDEs). Among them, three OCPs (i.e., hexachlorobenzene (HCB), p,p'-dichlorodiphenyltrichloroethane (p,p'-DDT), and p,p'-dichlorodiphenyldichloroethylene (p,p'-DDE)) and five PCBs (i.e., PCB52, PCB118, PCB138, PCB153, and PCB180) were detected in over 60% of the samples. PBDEs were not detected at a detection frequency of 60% or above. The most frequently detected POPs were p,p'-DDE (99.8%, geometric mean of 128.47 ng/g lipid), followed by PCB180 (98.8%, 8.49 ng/g lipid), PCB153 (98.8%, 13.14 ng/g lipid), HCB (96.2%, 67.08 ng/g lipid), PCB138 (95.2%, 8.84 ng/g lipid), PCB118 (89.6%, 2.66 ng/g lipid), p,p'-DDT (80.5%, 6.68 ng/g lipid), and PCB52 (71.2%, 1.57 ng/g lipid). The concentrations of most POPs were lower than or similar to concentrations reported in national-scale biomonitoring surveys. The only exception was HCB, whose concentration was up to seven-fold higher than the concentration reported by the Canadian Health Measures Survey. Excluding HCB and PCB52, most POPs showed increasing serum levels among older adults, adults with higher body mass index, adults living in coastal areas, and more frequent fish consumption. Relatively higher POP concentrations were observed in menopausal women. This study provides the first data on POP exposure levels among the representative adult population in Korea, and the results highlight the need to integrate POPs in the national biomonitoring program.

Crataegus pinnatifida Bunge Inhibits RANKL-Induced Osteoclast Differentiation in RAW 264.7 Cells and Prevents Bone Loss in an Ovariectomized Rat Model.

Osteoporosis is characterized by a decrease in bone microarchitecture with an increased risk of fracture. Long-term use of primary treatments, such as bisphosphonates and selective estrogen receptor modulators, results in various side effects. Therefore, it is necessary to develop alternative therapeutics derived from natural products. Crataegus pinnatifida Bunge (CPB) is a dried fruit used to treat diet-induced indigestion, loss of appetite, and diarrhea. However, research into the effects of CPB on osteoclast differentiation and osteoporosis is still limited. In vitro experiments were conducted to examine the effects of CPB on RANKL-induced osteoclast differentiation in RAW 264.7 cells. Moreover, we investigated the effects of CPB on bone loss in the femoral head in an ovariectomized rat model using microcomputed tomography. In vitro, tartrate-resistant acid phosphatase (TRAP) staining results showed the number of TRAP-positive cells, and TRAP activity significantly decreased following CPB treatment. CPB also significantly decreased pit formation. Furthermore, CPB inhibited osteoclast differentiation by suppressing NFATc1, and c-Fos expression. Moreover, CPB treatment inhibited osteoclast-related genes, such as Nfatc1, Ca2, Acp5, mmp9, CtsK, Oscar, and Atp6v0d2. In vivo, bone mineral density and structure model index were improved by administration of CPB. In conclusion, CPB prevented osteoclast differentiation in vitro and prevented bone loss in vivo. Therefore, CPB could be a potential alternative medicine for bone diseases, such as osteoporosis.

Anti-inflammatory effects of Fritillaria thunbergii Miquel extracts in LPS-stimulated murine macrophage RAW 264.7 cells.

The aim of the present study was to demonstrate that Fritillaria thunbergii Miquel extract exerts anti-inflammatory and antioxidant effects on lipopolysaccharide-stimulated RAW 264.7 cells. To confirm the inhibitory effect of ethyl acetate fraction of FTM (EAFM) on inflammation, the expression of nitric oxide (NO) and inflammatory cytokines was assessed by performing ELISA. Expression of intracellular mRNA and protein was confirmed by reverse transcription PCR and western blotting. In addition, the anti-inflammatory and anti-oxidant mechanisms of NF-κB, MAPK and heme oxygenase-1 (HO-1) were also investigated. EAFM significantly inhibited the expression of inflammatory factors including NO, IL-6 and TNF-α at non-toxic concentrations. EAFM also inhibited the mRNA and protein expression of inducible nitric oxide synthase in a concentration-dependent manner, but did not alter the expression of cyclooxygenase-2. Pre-treatment with EAFM inhibited the nuclear translocation of NF-κB, and suppressed the phosphorylation of ERK and JNK. In addition, EAFM induced 2,2'-azino-bis(3-ethylbenzothiazoline-6-sulfonic acid) radical scavenging activity and an increase in the expression of nuclear factor erythroid 2-related factor 2 (Nrf2) and HO-1. The results indicated that EAFM inhibited the expression of pro-inflammatory cytokines by inhibiting ERK/JNK phosphorylation and NF-κB translocation. EAFM also exerted antioxidant effects via Nrf2/HO-1 stimulation. Collectively, the results of the present study indicated that EAFM may be a valuable alternative for the treatment of a variety of inflammatory diseases.

Effects of Sparganii Rhizoma on Osteoclast Formation and Osteoblast Differentiation and on an OVX-Induced Bone Loss Model.

Postmenopausal osteoporosis is caused by an imbalance between osteoclasts and osteoblasts and causes severe bone loss. Osteoporotic medicines are classified into bone resorption inhibitors and bone formation promoters according to the mechanism of action. Long-term use of bisphosphonate and selective estrogen receptor modulators (SERMs) can cause severe side effects in postmenopausal osteoporosis patients. Therefore, it is important to find alternative natural products that reduce osteoclast activity and increase osteoblast formation. Sparganii Rhizoma (SR) is the dried tuberous rhizome of Sparganium stoloniferum Buchanan-Hamilton and is called "samreung" in Korea. However, to date, the effect of SR on osteoclast differentiation and the ovariectomized (OVX)-induced bone loss model has not been reported. In vitro, tartrate-resistant acid phosphatase (TRAP) staining, western blots, RT-PCR and other methods were used to examine the effect of SR on osteoclast differentiation and osteoblasts. In vivo, we confirmed the effect of SR in a model of OVX-induced postmenopausal osteoporosis. SR inhibited osteoclast differentiation and decreased the expression of TNF receptor-associated factor 6 (TRAF6), nuclear factor of activated T cells 1 (NFATc1) and c-Fos pathway. In addition, SR stimulates osteoblast differentiation and increased protein expression of the bone morphogenetic protein 2 (BMP-2)/SMAD signaling pathway. Moreover, SR protected against bone loss in OVX-induced rats. Our results appear to advance our knowledge of SR and successfully demonstrate its potential role as a osteoclastogenesis-inhibiting and osteogenesis-promoting herbal medicine for the treatment of postmenopausal osteoporosis.