Progranulin plays crucial roles in preserving bone mass by inhibiting TNF-α-induced osteoclastogenesis and promoting osteoblastic differentiation in mice.

A close correlation between atherosclerosis, inflammation, and osteoporosis has been recognized, although the precise mechanism remains unclear. The growth factor progranulin (PGRN) is expressed in various cells such as macrophages, leukocytes, and chondrocytes. PGRN plays critical roles in a variety of diseases, such as atherosclerosis and arthritis by inhibiting Tumor Necrosis Factor-α (TNF-α) signaling. The purpose of this study was to investigate the effect of PGRN on bone metabolism. Forty-eight-week old female homozygous PGRN knockout mice (PGRN-KO) (n = 8) demonstrated severe low bone mass in the distal femur compared to age- and sex-matched wild type C57BL/6J mice (WT) (n = 8) [BV/TV (%): 5.8 vs. 16.6; p < 0.001, trabecular number (1/mm): 1.6 vs. 3.8; p < 0.001]. In vitro, PGRN inhibited TNF-α-induced osteoclastogenesis from spleen cells of PGRN-KO mice. Moreover, PGRN significantly promoted ALP activity, osteoblast-related mRNA (ALP, osteocalcin) expression in a dose-dependent manner and up-regulated osteoblastic differentiation by down-regulating phosphorylation of ERK1/2 in mouse calvarial cells. In conclusion, PGRN may be a promising treatment target for both atherosclerosis and inflammation-related osteoporosis.

IL-6 negatively regulates osteoblast differentiation through the SHP2/MEK2 and SHP2/Akt2 pathways in vitro.

It has been suggested that interleukin-6 (IL-6)plays a key role in the pathogenesis of rheumatoid arthritis(RA), including osteoporosis not only in inflamed joints but also in the whole body. However, previous in vitro studies regarding the effects of IL-6 on osteoblast differentiation are inconsistent. The aim of this study was to examine the effects and signal transduction of IL-6 on osteoblast differentiation in MC3T3-E1 cells and primary murine calvarial osteoblasts. IL-6 and its soluble receptor significantly reduced alkaline phosphatase (ALP) activity, the expression of osteoblastic genes (Runx2, osterix, and osteocalcin), and mineralization in a dose-dependent manner, which indicates negative effects of IL-6 on osteoblast differentiation. Signal transduction studies demonstrated that IL-6 activated not only two major signaling pathways, SHP2/MEK/ERK and JAK/STAT3, but also the SHP2/PI3K/Akt2 signaling pathway. The negative effect of IL-6 on osteoblast differentiation was restored by inhibition of MEK as well as PI3K, while it was enhanced by inhibition of STAT3. Knockdown of MEK2 and Akt2 transfected with siRNA enhanced ALP activity and gene expression of Runx2. These results indicate that IL-6 negatively regulates osteoblast differentiation through SHP2/MEK2/ERK and SHP2/PI3K/Akt2 pathways, while affecting it positively through JAK/STAT3. Inhibition of MEK2 and Akt2 signaling in osteoblasts might be of potential use in the treatment of osteoporosis in RA.

Vitamin K2 administration is associated with decreased disease activity in patients with rheumatoid arthritis.

OBJECTIVES: Vitamin K2 (VitK2) is reported to induce not only bone mineralization of human osteoblasts and apoptosis of osteoclasts, but also apoptosis of rheumatoid arthritis (RA) synovial cells, while its clinical effect on disease activity of RA remains unknown. METHODS: 158 female RA patients (mean age 62.5 years) who had not been treated with warfarin, biologics, or teriparatide were enrolled in this study. VitK2 (45 mg/day) was administered in 70 patients with a serum undercarboxylated osteocalcin level of >4.5 ng/ml or with decreased bone mineral density in spite of the treatment with other anti-osteoporosis medications, regardless of RA disease activity. A longitudinal study was conducted in 52 patients who were additionally treated with VitK2 without changing their other medications for three months. RESULTS: In the cross-sectional study, as compared to the VitK2-naïve group (n = 88), the VitK2-treated group (n = 70) showed lower serum CRP (1.7 ± 0.2 vs. 0.5 ± 0.1 mg/dl; P < 0.001), MMP-3 (220.4 ± 21.9 vs. 118.0 ± 14.4 ng/ml; P < 0.001), and DAS28-CRP (2.9 ± 0.1 vs. 2.4 ± 0.1; P < 0.05). In the longitudinal study, patients who were additionally treated with VitK2 showed significant decreases in serum CRP (1.1 ± 0.2 to 0.6 ± 0.2 mg/dl; P < 0.001), MMP-3 (160.1 ± 25.6 to 125.0 ± 17.8 ng/ml; P < 0.05), and DAS28-CRP (3.1 ± 0.2 to 2.4 ± 0.1; P < 0.001). CONCLUSIONS: VitK2 may have the potential to improve disease activity besides osteoporosis in RA.

BMP-2/7 heterodimer strongly induces bone regeneration in the absence of increased soft tissue inflammation.

BACKGROUND CONTEXT: Bone morphogenetic protein (BMP)-2/7 heterodimer is a stronger inducer of bone regeneration than individual homodimers. However, clinical application of its potent bone induction ability may be hampered if its use is accompanied by excessive inflammatory reactions. PURPOSE: We sought to quantitatively evaluate bone induction and inflammatory reactions by BMP heterodimer and corresponding BMP homodimers using ultra-high resolution magnetic resonance imaging (MRI) and micro-computed tomography. STUDY DESIGN: An experimental animal study was carried out. METHODS: A total of 32 absorbable collagen sponge implantations into dorsal muscle were performed in rats of four different groups (control group, 0 µg BMP; recombinant human (rh)BMP-7 group, 3 µg rhBMP-7; rhBMP-2 group, 3 µg rhBMP-2; rhBMP-2/7 group, 3 µg rhBMP-2/7). Inflammatory reactions were evaluated by 11.7-T MRI (axial T2-weighted imaging using rapid acquisition with relaxation enhancement) at postoperative days 2 and 7. Bone volumes (BVs) of the induced ectopic bone were quantified at postoperative day 7. In addition, immunohistochemical staining for interleukin (IL)-1ß, IL-6, and tumor necrosis factor (TNF)-α was performed in samples obtained on postoperative day 2. Bone formation (BF)-to-inflammation (IM) ratios were calculated by dividing BVs by values of inflamed areas. RESULTS: At postoperative day 2, the mean volume of T2 high area on MRI scans in BMP-2 group was significantly larger than that in control group. In contrast, the BMP-2/7 had no difference in the mean volume of T2 high area compared with the control group; however, there was no difference between the BMP-2/7 compared with BMP-2 group. At postoperative day 7, the volumes of T2 high area were not different between the groups. Mean BV of the newly formed bone on postoperative day 7 was significantly greater in BMP-2/7 group than in BMP-7 groups. No new bone formation was observed in control group. BF-to-IM ratio in BMP-2/7 group was significantly higher than those in BMP-2 and BMP-7 homodimer groups. Immunohistochemistry experiments did not reveal differences in expression levels of IL-1ß, IL-6, or TNF-α in samples from BMP-2, BMP-7, and BMP-2/7 groups. CONCLUSIONS: This study demonstrated that BMP-2/7 heterodimer has stronger bone induction ability without accompanying increased inflammatory reactions (the increased BF-to-IM ratio) than those observed by BMP-2 or BMP-7 homodimers. These results suggest that BMP-2/7 heterodimer can be an alternative to BMP-2 and BMP-7 homodimers in clinical applications, although further translational studies, including whether lower doses of BMP heterodimer may produce similar bone formation compared with the BMP homodimers but produce a reduced inflammatory response, are required.

Comparison of the effects of forefoot joint-preserving arthroplasty and resection-replacement arthroplasty on walking plantar pressure distribution and patient-based outcomes in patients with rheumatoid arthritis.

PURPOSE: The purpose of this retrospective study is to clarify the difference in plantar pressure distribution during walking and related patient-based outcomes between forefoot joint-preserving arthroplasty and resection-replacement arthroplasty in patients with rheumatoid arthritis (RA). METHODS: Four groups of patients were recruited. Group1 included 22 feet of 11 healthy controls (age 48.6 years), Group2 included 36 feet of 28 RA patients with deformed non-operated feet (age 64.8 years, Disease activity score assessing 28 joints with CRP [DAS28-CRP] 2.3), Group3 included 27 feet of 20 RA patients with metatarsal head resection-replacement arthroplasty (age 60.7 years, post-operative duration 5.6 years, DAS28-CRP 2.4), and Group4 included 34 feet of 29 RA patients with metatarsophalangeal (MTP) joint-preserving arthroplasty (age 64.6 years, post-operative duration 3.2 years, DAS28-CRP 2.3). Patients were cross-sectionally examined by F-SCAN II to evaluate walking plantar pressure, and the self-administered foot evaluation questionnaire (SAFE-Q). Twenty joint-preserving arthroplasty feet were longitudinally examined at both pre- and post-operation. RESULTS: In the 1st MTP joint, Group4 showed higher pressure distribution (13.7%) than Group2 (8.0%) and Group3 (6.7%) (P<0.001). In the 2nd-3rd MTP joint, Group4 showed lower pressure distribution (9.0%) than Group2 (14.5%) (P<0.001) and Group3 (11.5%) (P<0.05). On longitudinal analysis, Group4 showed increased 1st MTP joint pressure (8.5% vs. 14.7%; P<0.001) and decreased 2nd-3rd MTP joint pressure (15.2% vs. 10.7%; P<0.01) distribution. In the SAFE-Q subscale scores, Group4 showed higher scores than Group3 in pain and pain-related scores (84.1 vs. 71.7; P<0.01) and in shoe-related scores (62.5 vs. 43.1; P<0.01). CONCLUSIONS: Joint-preserving arthroplasty resulted in higher 1st MTP joint and lower 2nd-3rd MTP joint pressures than resection-replacement arthroplasty, which were associated with better patient-based outcomes.

Modeling and remodeling effects of intermittent administration of teriparatide (parathyroid hormone 1-34) on bone morphogenetic protein-induced bone in a rat spinal fusion model.

BACKGROUND: Bone morphogenetic protein (BMP)-based tissue engineering has focused on inducing new bone efficiently. However, modeling and remodeling of BMP-induced bone have rarely been discussed. Teriparatide (parathyroid hormone [PTH] 1-34) administration initially increases markers of bone formation, followed by an increase in bone resorption markers. This unique activity would be expected to accelerate the modeling and remodeling of new BMP-induced bone. METHODS: Male Sprague-Dawley rats underwent posterolateral spinal fusion surgery and implantation of collagen sponge containing either 50 µg recombinant human (rh)BMP-2 or saline. PTH 1-34 (60 µg/kg, 3 times/week) or saline injections were continued from preoperative week 2 week to postoperative week 12. The volume and quality of newly formed bone were monitored by in vivo micro-computed tomography and analyses of bone histomorphometry and serum bone metabolism markers were conducted at postoperative week 12. RESULTS: Microstructural indices of the newly formed bone were significantly improved by PTH 1-34 administration, which significantly decreased the tissue volumes of the fusion mass at postoperative week 12 compared to that at postoperative week 2. Bone histomorphometry and serum analyses showed that PTH administration significantly increased both bone formation and resorption markers. Analysis of the histomorphometry of cortical bone identified predominant periosteal bone resorption and endosteal bone formation. CONCLUSIONS: Long-term intermittent administration of PTH 1-34 significantly accelerated the modeling and remodeling of new BMP-induced bone. CLINICAL RELEVANCE: Our results suggest that the combined administration of rhBMP-2 and PTH 1-34 facilitates qualitative and quantitative improvements in bone regeneration, by accelerating bone modeling and remodeling.

Intermittent administration of teriparatide enhances graft bone healing and accelerates spinal fusion in rats with glucocorticoid-induced osteoporosis.

BACKGROUND CONTEXT: There has been no study regarding the effect of intermittent administration of teriparatide (TPTD [recombinant human parathyroid hormone (1-34)]) on spinal fusion in patients with glucocorticoid-induced osteoporosis (GIOP). PURPOSE: To elucidate the effect of intermittent administration of TPTD on spinal fusion in rats with GIOP. STUDY DESIGN: An experimental animal study of rats under continuous glucocorticoid (GC) exposure undergoing spinal fusion surgery and administration of TPTD or saline. METHODS: Male 8-week-old rats (n=18) were administered 5 mg/kg methylprednisolone (MP) for 12 weeks. After 6 weeks of MP administration, the rats underwent posterolateral spinal fusion (L4-L5) with iliac crest autograft. Then, five times a week, they were given either saline or 40 µg/kg TPTD for 6 weeks. The following assessments were performed: time-course bone microstructural analysis of the fusion mass and adjacent vertebrae (L6), with in vivo microcomputed tomography (µCT); fusion assessment, with manual palpation testing and three-dimensional CT images; and bone histomorphometrical analysis of the fusion mass. RESULTS: In the TPTD group, values for bone volume and other bone microstructural parameters at the fusion mass increased and peaked 4 weeks after surgery, and these values were significantly greater than those for the control (CNT) group at 4 and 6 weeks after surgery. Fusion assessment showed that fusion rate was higher in the TPTD group than in the CNT group (CNT group: 56%, TPTD group: 89%). Bone histomorphometry revealed that values for bone formation parameters were significantly higher in the TPTD group than in the CNT group. CONCLUSIONS: Under continuous GC exposure in a rat model of spinal fusion, intermittent TPTD administration accelerated bone modeling and remodeling predominantly by stimulating bone formation at the fusion mass and increasing the fusion rate. Intermittent TPTD administration also improved bone microarchitecture of adjacent vertebrae.

The bone morphogenetic protein-2/7 heterodimer is a stronger inducer of bone regeneration than the individual homodimers in a rat spinal fusion model.

BACKGROUND CONTEXT: Bone morphogenetic proteins (BMPs) are a group of dimeric growth factors that belong to the transforming growth factor super family and are capable of eliciting new bone formation. Previous studies have suggested that the coexpression of two different BMP genes in a cell can result in the production of BMP heterodimers that are more potent than homodimers. However, because of the difficulty in optimizing the level of BMP gene expression, the coexpression of two different BMP genes also produces BMP homodimers as a by-product. These homodimers could, in theory, interact with the heterodimers. PURPOSE: To elucidate the effects of a BMP-2/7 heterodimer, which were investigated in depth using purified BMP-2/7 heterodimers, BMP-2 homodimers, and BMP-7 homodimers in a rat spinal fusion model. METHODS: Bilateral posterolateral fusion at L4-L5 was performed in four different groups: control group animals were implanted with collagen carriers alone; BMP-7 group animals with collagen carriers+1 µg of BMP-7 homodimer; BMP-2 group animals with collagen carriers+1 µg of BMP-2 homodimer; and BMP-2/7 group animals with collagen carriers+1 µg of the BMP-2/7 heterodimer. The following assessments were performed: bone microstructural analysis of the fusion mass and tissue volume (TV) with microcomputed tomography (micro-CT); fusion assessment with manual palpation testing and three-dimensional CT images; and bone histomorphometrical analysis of the fusion mass. RESULTS: The fusion scores, as determined by radiography, and the TV of the newly formed bone, as determined by micro-CT, were significantly higher in the BMP-2/7 heterodimer group than the other groups (p<.0001). The microstructural indices of the newly formed bone did not differ between the groups. Moreover, histologic analysis of the fused spines revealed that the formation of the trabecular bone bridging the transverse process was the highest in this group. CONCLUSIONS: This study demonstrated that BMP-2/7 heterodimer is a stronger inducer of bone regeneration than BMP-2 or -7 homodimers. The use of a purified BMP-2/7 heterodimer may represent an efficient alternative to the current clinical use of BMP-2 or -7 homodimers. Further studies as to the side effects of BMP-2/7 heterodimer are required.

Effect of Intermittent Administration of Teriparatide (Parathyroid Hormone 1-34) on Bone Morphogenetic Protein-Induced Bone Formation in a Rat Model of Spinal Fusion.

BACKGROUND: Although clinical bone morphogenetic protein (BMP) therapy is effective at enhancing bone formation in patients managed with spinal arthrodesis, the required doses are very high. Teriparatide (parathyroid hormone 1-34) is approved by the U.S. Food and Drug Administration to treat osteoporosis and is a potent anabolic agent. In this study, intermittent administration of parathyroid hormone 1-34 combined with transplantation of BMP was performed to elucidate the effect of parathyroid hormone 1-34 on the fusion rate and quality of newly formed bone in a rat model. METHODS: A total of forty-eight male Sprague-Dawley rats underwent posterolateral lumbar spinal arthrodesis with one of three different treatments with recombinant human (rh) BMP-2: (1) 0 µg (control), (2) 2 µg (low dose), or (3) 50 µg (high dose). Each of the rhBMP-2 treatments was studied in combination with intermittent injections of either parathyroid hormone 1-34 (180 µg/kg/wk) or saline solution starting two weeks before the operation and continuing until six weeks after the operation. Osseous fusion was assessed with use of radiographs and a manual palpation test. Microstructural indices of the newly formed bone were evaluated with use of micro-computed tomography. The serum markers of bone metabolism were also quantified. RESULTS: The fusion rate in the group treated with 2 µg of rhBMP-2 significantly increased (from 57% to 100%) with the administration of parathyroid hormone 1-34 (p < 0.05). The fusion rates in the other groups did not change significantly with the administration of parathyroid hormone 1-34. The bone volume density of the newly formed bone significantly increased in both the 2-µg and 50-µg rhBMP-2 treatment groups with the administration of parathyroid hormone 1-34 (p < 0.01). Micro-computed tomography scans of the newly formed bone clearly demonstrated an abundance of trabecular bone formation in the group treated with parathyroid hormone 1-34. In addition, serum levels of osteocalcin were significantly increased in the parathyroid hormone 1-34 treatment group. CONCLUSIONS: Intermittent administration of parathyroid hormone 1-34 significantly increased fusion rates in the group treated with low-dose rhBMP-2, and it improved the quality of the newly formed bone in both the high and low-dose groups in a rat model of rhBMP-2-induced spinal fusion. CLINICAL RELEVANCE: Our results suggest that the combined administration of rhBMP-2 and parathyroid hormone 1-34 may lead to efficient bone regeneration.

Malignant phosphaturic mesenchymal tumor of the pelvis: A report of two cases.

Tumor-induced osteomalacia (TIO) is a rare acquired form of hypophosphatemia commonly associated with phosphaturic mesenchymal tumors (PMTs) located in the bone or soft tissue. Resection of the tumor can cure osteomalacia. Fibroblast growth factor 23 has been identified as a major pathophysiological factor responsible for phosphaturia. The majority of PMTs are benign, and malignant PMTs are uncommon. Even in rare cases, the malignant transformation of PMTs is extremely uncommon. The current study presents two cases in which the patients succumbed to malignant PMTs that developed in the pelvis. The first patient was a 35-year-old female with a malignant PMT occurring as a synchronous double cancer associated with papillary thyroid carcinoma. Diagnosis was difficult, as the multiple uptake on positron emission tomography with 18F-fluorodeoxyglucose presented as pseudofractures mimicking the metastases of thyroid carcinoma. The patient succumbed to rapidly progressive lung metastases. The second patient presented with a pelvic tumor that had developed over 26 years. The patient was diagnosed with a benign PMT by open biopsy and a complete resection was performed. However, two years later, the tumor recurred and lung metastases were observed. The patient ultimately succumbed to respiratory failure due to relapsing lung metastases and disseminated intravascular coagulation. These two cases demonstrate the potential lethality of malignant PMTs and the malignant transformation of benign PMTs. Therefore, TIO patients must be followed up even if diagnosed with a benign tumor. Although TIO is an extremely rare disease, the possibility of malignant PMTs must be recognized.

Oxygen and air nanobubble water solution promote the growth of plants, fishes, and mice.

Nanobubbles (<200 nm in diameter) have several unique properties such as long lifetime in liquid owing to its negatively charged surface, and its high gas solubility into the liquid owing to its high internal pressure. They are used in variety of fields including diagnostic aids and drug delivery, while there are no reports assessing their effects on the growth of lives. Nanobubbles of air or oxygen gas were generated using a nanobubble aerator (BUVITAS; Ligaric Company Limited, Osaka, Japan). Brassica campestris were cultured hydroponically for 4 weeks within air-nanobubble water or within normal water. Sweetfish (for 3 weeks) and rainbow trout (for 6 weeks) were kept either within air-nanobubble water or within normal water. Finally, 5 week-old male DBA1/J mice were bred with normal free-chaw and free-drinking either of oxygen-nanobubble water or of normal water for 12 weeks. Oxygen-nanobubble significantly increased the dissolved oxygen concentration of water as well as concentration/size of nanobubbles which were relatively stable for 70 days. Air-nanobubble water significantly promoted the height (19.1 vs. 16.7 cm; P<0.05), length of leaves (24.4 vs. 22.4 cm; P<0.01), and aerial fresh weight (27.3 vs. 20.3 g; P<0.01) of Brassica campestris compared to normal water. Total weight of sweetfish increased from 3.0 to 6.4 kg in normal water, whereas it increased from 3.0 to 10.2 kg in air-nanobubble water. In addition, total weight of rainbow trout increased from 50.0 to 129.5 kg in normal water, whereas it increased from 50.0 to 148.0 kg in air-nanobubble water. Free oral intake of oxygen-nanobubble water significantly promoted the weight (23.5 vs. 21.8 g; P<0.01) and the length (17.0 vs. 16.1 cm; P<0.001) of mice compared to that of normal water. We have demonstrated for the first time that oxygen and air-nanobubble water may be potentially effective tools for the growth of lives.