Abnormal kynurenine level contributes to the pathological bone features of ankylosing spondylitis.

OBJECTIVE: Ankylosing spondylitis (AS) exhibits paradoxical bone features typically characterized by new bone formation and systemic bone loss. Although abnormal kynurenine (Kyn), a tryptophan metabolite, has been closely linked to the disease activity of AS, the distinct role of its pathological bone features remains unknown. METHODS: Kynurenine sera level was collected from healthy control (HC; n = 22) and AS (n = 87) patients and measured by ELISA. In the AS group, we analyzed and compared the Kyn level based on the modified stoke ankylosing spondylitis spinal score (mSASSS), MMP13, and OCN. Under osteoblast differentiation, the treatment with Kyn in AS-osteoprogenitors conducted cell proliferation, alkaline phosphatase activity, bone mineralization-related alizarin red s (ARS), von kossa (VON), hydroxyapatite (HA) staining, and mRNA expression markers (ALP, RUNX2, OCN, and OPG) for bone formation. TRAP and F-actin staining was used for osteoclast formation of mouse osteoclast precursors. RESULTS: Kyn sera level was significantly elevated in the AS group compared to the HC. In addition, Kyn sera level was correlated with mSASSS (r = 0.03888, p = 0.067), MMP13 (r = 0.0327, p = 0.093), and OCN (r = 0.0436, p = 0.052). During osteoblast differentiation, treatment with Kyn exhibited no difference in cell proliferation and alkaline phosphate (ALP) activity for bone matrix maturation but promoted ARS, VON, and HA staining for bone mineralization. Interestingly, osteoprotegerin (OPG) and OCN expressions of AS-osteoprogenitors were augmented in the Kyn treatment during differentiation. In growth medium, Kyn treatment of AS-osteoprogenitors resulted in induction of OPG mRNA, protein expression, and Kyn-response genes (AhRR, CYP1b1, and TIPARP). Secreted OPG proteins were observed in the supernatant of AS-osteoprogenitors treated with Kyn. Notably, the supernatant of Kyn-treated AS-osteoprogenitors interrupted the RANKL-mediated osteoclastogenesis of mouse osteoclast precursor such as TRAP-positive osteoclast formation, NFATc1 expression, and osteoclast differentiation markers. CONCLUSION: Our results revealed that elevated Kyn level increased the bone mineralization of osteoblast differentiation in AS and decreased RANKL-mediated osteoclast differentiation by inducing OPG expression. Out study have implication for potential coupling factors linking osteoclast and osteoblast where abnormal Kyn level could be involved in pathological bone features of AS.

Dikkopf-1 promotes matrix mineralization of osteoblasts by regulating Ca+-CAMK2A- CREB1 pathway.

Dickkopf-1 (DKK1) is a secreted protein that acts as an antagonist of the canonical WNT/ß-catenin pathway, which regulates osteoblast differentiation. However, the role of DKK1 on osteoblast differentiation has not yet been fully clarified. Here, we investigate the functional role of DKK1 on osteoblast differentiation. Primary osteoprogenitor cells were isolated from human spinal bone tissues. To examine the role of DKK1 in osteoblast differentiation, we manipulated the expression of DKK1, and the cells were differentiated into mature osteoblasts. DKK1 overexpression in osteoprogenitor cells promoted matrix mineralization of osteoblast differentiation but did not promote matrix maturation. DKK1 increased Ca+ influx and activation of the Ca+/calmodulin-dependent protein kinase II Alpha (CAMK2A)-cAMP response element-binding protein 1 (CREB1) and increased translocation of p-CREB1 into the nucleus. In contrast, stable DKK1 knockdown in human osteosarcoma cell line SaOS2 exhibited reduced nuclear translocation of p-CREB1 and matrix mineralization. Overall, we suggest that manipulating DKK1 regulates the matrix mineralization of osteoblasts by Ca+-CAMK2A-CREB1, and DKK1 is a crucial gene for bone mineralization of osteoblasts. [BMB Reports 2022; 55(12): 627-632].

Blocking TNFα attenuates progressive cartilage matrix degradation in inflammatory arthritis.

Because damage to hyaline cartilage is irreversible, relieving progressive cartilage destruction is an important therapeutic approach for inflammatory arthritis. In the present study, human hyaline chondrocytes were isolated from total knee replacements of 15 patients with osteoarthritis (OA) and three with rheumatoid arthritis (RA). Synovial fluid of OA (n=25) and RA (n=34) were collected to measure tumor necrosis factor α (TNFα) using ELISA. Consistent with previous studies, the synovial fluid exhibited high TNFα levels and hyaline cartilage was severely destroyed in patients with RA. TNFα-treated chondrocytes were used as model for inflammatory arthritis. TNFα did not influence proliferation or extracellular matrix expression in chondrocytes, but induced matrix metalloproteinase (MMP)1, 3 and 13 expression levels in chondrocytes, which was accompanied by activation of nuclear factor-κB signaling. During chondrogenic differentiation, TNFα attenuated mRNA expression levels of anabolic factors (collagen type 2 and aggrecan) and enhanced mRNA expression of catabolic factors (MMP1, MMP3 and MMP13) in chondrocytes. Moreover, anti-TNFα agents (Golimumab) inhibited the TNFα-induced metabolic shift in chondrocytes and chondrogenic differentiation. The present study revealed a mechanism by which TNFα may induce metabolic shift in chondrocytes, leading to progressive chondrocyte destruction.

The TNF-NF-kB-DKK1 Axis Promoted Bone Formation in the Enthesis of Ankylosing Spondylitis.

Objective: This study aimed to determine the serum Dickkopf 1 (DKK1) levels in ankylosing spondylitis (AS) patients and decipher the mechanism of tumor necrosis factor (TNF)-mediated DKK1 regulation in human AS enthesis cells. Methods: The sera were obtained from 103 patients with AS and 30 healthy controls (HCs) The enthesis of facet joints were obtained from 4 AS patients and 5 controls The serum levels of DKK1 were measured using ELISA and compared between AS and HCs The impact of TNF on DKK1 expression in human primary spinal enthesis cells was evaluated using various molecular biology techniques and bone formation indicators. Results: AS patients showed higher serum DKK1 levels than HCs after adjusting for age (9174 [6153∼1,3100] pg/mL vs 8262 [6703∼9278] pg/mL, p=0043) TNF treatment promoted bone formation and DKK1 expression in both control enthesis cells and those of AS This enhanced bone formation by TNF was pronounced in AS-enthesis than those of controls Mechanically, TNF induced NF-kB activation upregulates the DKK1 transcript level While, NF-kB inhibitor led to downregulate DKK1 expression in the enthesis Besides, DKK1 overexpression promoted bone formation in enthesis. Conclusion: TNF induced DKK1 expression in the enthesis through NF-kB activation TNF-induced DKK1 expression may play a bone formation in the radiologic progression of ankylosing spondylitis.

TGFß1 Suppressed Matrix Mineralization of Osteoblasts Differentiation by Regulating SMURF1-C/EBPß-DKK1 Axis.

Transforming growth factor ß1 (TGFß1) is a major mediator in the modulation of osteoblast differentiation. However, the underlying molecular mechanism is still not fully understood. Here, we show that TGFß1 has a dual stage-dependent role in osteoblast differentiation; TGFß1 induced matrix maturation but inhibited matrix mineralization. We discovered the underlying mechanism of the TGFß1 inhibitory role in mineralization using human osteoprogenitors. In particular, the matrix mineralization-related genes of osteoblasts such as osteocalcin (OCN), Dickkopf 1 (DKK1), and CCAAT/enhancer-binding protein beta (C/EBPß) were dramatically suppressed by TGFß1 treatment. The suppressive effects of TGFß1 were reversed with anti-TGFß1 treatment. Mechanically, TGFß1 decreased protein levels of C/EBPß without changing mRNA levels and reduced both mRNA and protein levels of DKK1. The degradation of the C/EBPß protein by TGFß1 was dependent on the ubiquitin-proteasome pathway. TGFß1 degraded the C/EBPß protein by inducing the expression of the E3 ubiquitin ligase Smad ubiquitin regulatory factor 1 (SMURF1) at the transcript level, thereby reducing the C/EBPß-DKK1 regulatory mechanism. Collectively, our findings suggest that TGFß1 suppressed the matrix mineralization of osteoblast differentiation by regulating the SMURF1-C/EBPß-DKK1 axis.

Anterior cruciate ligament remnant cells have different potentials for cell differentiation based on their location.

Histological and cytological observations of the human anterior cruciate ligament (ACL) had been described, but the differentiation potency based on their location is still unknown. To determine and compare proliferation and differentiation potential of cells derived from distal and middle thirds of the ACL remnant, ACL remnant was initially marked at the distal third (within 10 mm from the tibial insertion) and middle third (between 10-20 mm from the tibial insertion) and then dissected. Both the middle and distal third regions of ACL remnant were analyzed using CD34+ cell counting. Cell proliferation rate did not differ in both middle and distal third regions of ACL remnant, but they showed different characteristics in cell differentiation depending on their location. The distal third region of the ACL remnant had a tendency for chondrogenic differentiation with higher expression of CD34+ cells. On the other hand, the middle third region of ACL remnant had a strong tendency for osteogenic and ligamentous differentiation. Characteristics of the ACL remnant tissues should be considered when performing remnant-preserving or harvesting ACL remnants for tissue engineering.

DKK1 Induced by 1,25D3 Is Required for the Mineralization of Osteoblasts.

1α,25-dihydroxyvitamin D3 (1,25D3), the most popular drug for osteoporosis treatment, drives osteoblast differentiation and bone mineralization. Wnt/ß-catenin signaling is involved in commitment and differentiation of osteoblasts, but the role of the Dickkopf-related protein 1 (DKK1), a Wnt antagonist, in osteoblasts remains unknown. Here, we demonstrate the molecular mechanism of DKK1 induction by 1,25D3 and its physiological role during osteoblast differentiation. 1,25D3 markedly promoted the expression of both CCAAT/enhancer binding protein beta (C/EBPß) and DKK1 at day 7 during osteoblast differentiation. Interestingly, mRNA and protein levels of C/EBPß and DKK1 in osteoblasts were elevated by 1,25D3. We also found that C/EBPß, in response to 1,25D3, directly binds to the human DKK1 promoter. Knockdown of C/EBPß downregulated the expression of DKK1 in osteoblasts, which was partially reversed by 1,25D3. In contrast, overexpression of C/EBPß upregulated DKK1 expression in osteoblasts, which was enhanced by 1,25D3. Furthermore, 1,25D3 treatment in osteoblasts stimulated secretion of DKK1 protein within the endoplasmic reticulum to extracellular. Intriguingly, blocking DKK1 attenuated calcified nodule formation in mineralized osteoblasts, but not ALP activity or collagen synthesis. Taken together, these observations suggest that 1,25D3 promotes the mineralization of osteoblasts through activation of DKK1 followed by an increase of C/EBPß.

Physiological Factor Evaluation of the Warm Humidification of Anesthetic Gas Nursing Care.

OBJECTIVE: This study aimed to evaluate the effects of warm humidification of anesthetic gas nursing care on physiological factors in elderly patients during laparoscopic cancer surgery. METHODS: Sixty elderly laparoscopic surgical patients were recruited from a medical center in Korea. The participants' physiological factors (blood pressure, body temperature, and leukocyte and neutrophil counts) were measured four times. RESULTS: After warm humidification of anesthetic gas nursing care, there were significant differences in body temperatures. CONCLUSIONS: These findings indicate that warm humidification of anesthetic gas nursing care is effective in maintaining body temperature. Thus, this type of nursing care can be effective in elderly cancer patients undergoing laparoscopic surgery.

Low-temperature synthesis of LiFePO4 nanocrystals by solvothermal route.

LiFePO4 nanocrystals were synthesized at a very low temperature of 170°C using carbon nanoparticles by a solvothermal process in a polyol medium, namely diethylene glycol without any heat treatment as a post procedure. The powder X-ray diffraction pattern of the LiFePO4 was indexed well to a pure orthorhombic system of olivine structure (space group: Pnma) with no undesirable impurities. The LiFePO4 nanocrystals synthesized at low temperature exhibited mono-dispersed and carbon-mixed plate-type LiFePO4 nanoparticles with average length, width, and thickness of approximately 100 to 300 nm, 100 to 200 nm, and 50 nm, respectively. It also appeared to reveal considerably enhanced electrochemical properties when compared to those of pristine LiFePO4. These observed results clearly indicate the effect of carbon in improving the reactivity and synthesis of LiFePO4 nanoparticles at a significantly lower temperature.

The effects of Mo doping on 0.3Li[Li0.33Mn0.67]O2·0.7Li[Ni0.5Co0.2Mn0.3]O2 cathode material.

Mo doped Li excess transition metal oxides formulated as 0.3Li[Li(0.33)Mn(0.67)]O(2)·0.7Li[Ni(0.5-x)Co(0.2)Mn(0.3-x)Mo(2x)]O(2) were synthesized using the co-precipitation process. The effects of the substitution of Ni and Mn with Mo were investigated for the density of the states, the structure, cycling stability, rate performance and thermal stability by tools such as first principle calculations, synchrotron X-ray diffraction, field-emission SEM, solid state (7)Li MAS nuclear magnetic resonance (NMR), X-ray photoelectron spectroscopy (XPS), elemental mapping by scanning TEM (STEM), inductively coupled plasma atomic emission spectrometry (ICP-AES) and a differential scanning calorimeter (DSC). It was confirmed that high valence Mo(6+) doping of the Li-excess manganese-nickel-cobalt layered oxide in the transition metal enhanced the structural stability and electrochemical performance. This increase was due to strong Mo-O hybridization inducing weak Ni-O hybridization, which may reduce O(2) evolution, and metallic behavior resulting in a diminishing cell resistance.

Synthesis and characterization of integrated layered nanocomposites for lithium ion batteries.

The series of Li[NixMxLi1/3-xMn2/3-x]O2 cathodes, where M is cobalt or chromium with a wide compositional range x from 0 to 0.33, were prepared by hydroxide coprecipitation method with subsequent quenching. The sample structures were investigated using X-ray diffraction results which were indexed completely on the basis of a trigonal structure of space group R3m̄ with monoclinic C2/m phase as expected. The morphologies and electrochemical properties of the samples obtained were compared as the value of x and substituted transition metal. The particle sizes of cobalt-substituted Li[NixCoxLi1/3-xMn2/3-x]O2 samples are much smaller than those of the Li[NixCrxLi1/3-xMn2/3-x]O2 system. The electrode containing Li[NixCoxLi1/3-xMn2/3-x]O2 with x = 0.10 delivered a discharge capacity of above 200 mAh/g after 10 cycles due to the activation of Li2MnO3.PACS: 82.47.Aa; 82.47.-a; 82.45.Fk.

[A comparison of the effects of intravenous fluid warming and skin surface warming on peri-operative body temperature and acid base balance of elderly patients with abdominal surgery].

PURPOSE: The purpose of this study was to compare the effects of intravenous fluid warming and skin surface warming on peri-operative body temperature and acid base balance of abdominal surgical patients under general anesthesia. METHOD: Data collection was performed from January 4th, to May 31, 2004. The intravenous fluid warming(IFW) group (30 elderly patients) was warmed through an IV line by an Animec set to 37 degrees C. The skin surface warming (SSW) group (30 elderly patients) was warmed by a circulating-water blanket set to 38 degrees C under the back and a 60W heating lamp 40 cm above the chest. The warming continued from induction of general anesthesia to two hours after completion of surgery. Collected data was analyzed using Repeated Measures ANOVA, and Bonferroni methods. RESULTS: SSW was more effective than IFW in preventing hypothermia(p= .043), preventing a decrease of HCO(3)(-)(p= .000) and preventing base excess (p= .000) respectively. However, there was no difference in pH between the SSW and IFW (p= .401) groups. CONCLUSION: We conclude that skin surface warming is more effective in preventing hypothermia, and HCO(3)(-) and base excess during general anesthesia, and returning to normal body temperature after surgery than intravenous fluid warming; however, skin surface warming wasn't able to sustain a normal body temperature in elderly patients undergoing abdominal surgery under general anesthesia.

Molecular characterization of PL97-1, the first Korean isolate of the porcine reproductive and respiratory syndrome virus.

We determined the complete nucleotide and predicted amino acid sequence of the genomic RNA of PL97-1, the first Korean strain of porcine reproductive and respiratory syndrome virus (PRRSV), which was isolated from the serum of an infected pig in 1997. We found that the 15411-nucleotide genome of PL97-1 consisted of a 189-nucleotide 5' noncoding region (NCR), a 15071-nucleotide protein-coding region, and a 151-nucleotide 3'NCR, followed by a poly (A) tail. The 5'-end of PL97-1 began with 1ATG ACG TAT AGG12. Comparison of the PL97-1 genome with the 11 fully sequenced PRRSV genomes currently available revealed sequence divergence ranging from 0.3% (the VR-2332-derived vaccine MLV RespPRRS/Repro strain) to 38% (the Dutch Lelystad strain). To better understand the genetic relationships between these different strains, phylogenetic analyses were performed on the full-length PRRSV genomes. Significantly, the phylogenetic tree based on the ORF1b or ORF7 genes most closely resembled the tree based on the full-length genomes. Thus, these single genes will be the most useful in revealing the genetic relationships between the different strains relative to their geographical distribution. Extensive phylogenetic analyses using the ORF7 sequences of 111 PRRSV isolates available revealed that PL97-1 is most closely related to the North American genotype VR-2332, a VR-2332-derived vaccine strain, and Chinese BJ-4. It is distantly related to the European genotype Lelystad. This study provides the largest full-length genome phylogenetic analysis of PRRSV that has been published to date, and supports an earlier genetic grouping of the many temporally and geographically diverse PRRSV strains currently isolated.