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Yuichi Ono, Naohisa Miyakoshi, Yuji Kasukawa, Manabu Akagawa, Ryota Kimura, Itsuki Nagahata, Yusuke Yuasa, Chiaki Sato, Yoichi Shimada Department of Orthopedic Surgery, Akita University Graduate School of Medicine, Akita, JapanCorrespondence: Yuichi OnoDepartment of Orthopedic Surgery, Akita University Graduate School of Medicine, 1-1-1, Hondo, Akita-shi, Akita 010-8543, JapanTel +81-18-884-6148Fax +81-18-836-2617Email yuichi.ono24@gmail.comPurpose: Sarcopenia and osteoporosis are both serious health problems in postmenopausal women. The Asia Working Group for Sarcopenia recommends using the skeletal muscle index (SMI), which is height-adjusted appendicular skeletal muscle mass (ASMM). However, loss of height has been shown to be a common clinical finding in patients with osteoporosis. This study examined the prevalence of presarcopenia using height and arm span, which is a predictor of height, and investigated the diagnostic accuracy for presarcopenia.Methods: A total of 55 post-menopausal osteoporotic patients aged 62– 95 years underwent bioelectrical impedance analysis (BIA) for ASMM measurement and dual-energy X-ray absorptiometry (DXA) scan for bone mineral density (BMD). Anthropometric measurements, including height, weight, and arm span were taken, and body mass index (BMI), SMI, and arm span-adjusted SMI (Arm span SMI) were calculated. Presarcopenia was defined as SMI or Arm span SMI < 5.7 kg/m2 in this study.Results: The prevalence of presarcopenia was 27.3% and 38.2% evaluated by SMI and Arm span SMI, respectively. The prevalence of presarcopenia was higher when evaluated by Arm span SMI than by SMI. In the presarcopenia group diagnosed only by Arm span SMI (n=11), the arm span-height difference was significantly higher (p< 0.001) and the percentage of young adult mean (YAM) femoral neck-BMD was significantly lower (p=0.013) compared to the normal group diagnosed by both SMI and Arm span-SMI (n=29).Conclusion: These results indicated that Arm span SMI might be useful for the diagnosis of sarcopenia in patients with severe osteoporosis and kyphosis.Keywords: arm span, body height, osteoporosis, presarcopenia, skeletal muscle mass

β-Caryophyllene (BCP), a dietary phytocannabinoid, significantly suppresses palmitate-induced lipid accumulation in human HepG2 hepatocytes via activation of AMP-activated protein kinase (AMPK) signaling. The objective of the preset research was to assess whether oral administration of BCP alleviates obesity-induced hepatic steatosis in mice through AMPK activation. We examined the protective action of supplementation of 0.3% BCP (w/w) in a high-fat diet (HFD) on C57BL/6J mice for 12 weeks. BCP supplementation evidently ameliorated histological hepatic steatosis features, and significantly reduced triglycerides and cholesterol levels in the liver, and serum levels of aspartate aminotransferase and alanine aminotransferase as compared with non-supplemented HFD-fed mice. Immunoblotting revealed that BCP supplementation in HFD-fed mice also caused hepatic AMPK activation. Furthermore, treatment with BCP in HFD-fed mice significantly suppressed body weight gain and attenuated obesity-related phenotypes relative to the HFD mice. Our results suggest the usefulness of BCP in the prevention of obesity-related liver steatosis and liver injury., (© The Author(s) 2024. Published by Oxford University Press on behalf of Japan Society for Bioscience, Biotechnology, and Agrochemistry.)

Optimal ligament balance is crucial for successful total knee arthroplasty (TKA), which is a standard procedure for managing end-stage knee osteoarthritis. However, the reported values of ligament balance vary due to different methodologies and population differences, which requires further studies. This study aimed to investigate the varus-valgus ligament balance in healthy knees of Japanese individuals and its relationship with lower limb alignment. The studyanalyzed64 knees from 33 healthy young adults using stress radiographs to measure varus-valgus laxity. The mechanical hip-knee-ankle angle, medial proximal tibial angle, and lateral distal femoral angles were determined using digital long-leg radiographs. Varus and valgus laxities were compared, and the relationship between ligament balance and alignment parameters was analyzed. Mean varus laxity (3.6°± 1.3°) was significantly greater than mean valgus laxity(2.9°± 1.0°) (p<0.0001). No significant association was observed between ligament balance and alignment parameters. Healthy knees of Japanese individuals exhibited slightly greater varus laxity (0.7°) than valgus laxity, with no correlation observed between ligament balance and lower limb alignment. The finding underscores the importance of recognizing alignment diversity while noting that alignment does not influence ligament balance; this is particularly relevant in modern TKA techniques focusing on patient-specific joint and ligament reconstructions. The study's findings could help formulate strategies benefiting knee osteoarthritis management., Competing Interests: The authors have declared that no competing interests exist., (Copyright: © 2024 Akagawa et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.)

Although several studies have suggested a possible association between sarcopenia and knee osteoarthritis (OA) in the elderly, there remains no definitive evidence. Recently, however, the serum creatinine/cystatin C ratio (sarcopenia index: SI) was reported to correlate with skeletal muscle mass. The present retrospective study therefore investigated the impact of reduced skeletal muscle mass on advanced knee OA using SI. In 55 individuals scheduled for knee osteotomy or knee arthroplasty, correlations between SI and patient-reported outcomes such as the Knee Society Score (KSS), Knee Injury and Osteoarthritis Outcome Score (KOOS), and Oxford Knee Score (OKS) were explored. Significant associations were found between SI and the KSS functional activity score (β=0.37; p=0.022), KOOS subscale for activities of daily living (β=0.42; p=0.0096), and OKS (β=0.42; p=0.0095). This study underscores the role of reduced muscle mass in functional outcomes and introduces SI as a valuable marker for assessing muscle loss in knee OA patients., Competing Interests: No potential conflict of interest relevant to this article was reported.

Pyrroloquinoline quinone disodium salt (PQQ) is a red trihydrate crystal that was approved as a new food ingredient by FDA in 2008. Now, it is approved as a food in Japan and the EU. PQQ has redox properties and exerts antioxidant, neuroprotective, and mitochondrial biogenesis effects. The baseline intake level of PQQ is considered to be 20 mg/day. PQQ ingestion lowers blood lipid peroxide levels in humans, suggesting antioxidant activity. In the field of cognitive function, double-blind, placebo-controlled trials have been conducted. Various improvements have been reported regarding general memory, verbal memory, working memory, and attention. Furthermore, a stratified analysis of a population with a wide range of ages revealed unique effects in young people (20-40 years old) that were not observed in older adults (41-65 years old). Specifically, cognitive flexibility and executive speed improved more rapidly in young people at 8 weeks. Co-administration of PQQ and coenzyme Q10 further enhanced these effects. In an open-label trial, PQQ was shown to improve sleep and mood. Additionally, PQQ was found to suppress skin moisture loss and increase PGC-1α expression. Overall, PQQ is a food with various functions, including brain health benefits. J. Med. Invest. 71 : 23-28, February, 2024.

Objectives: Atypical femoral fracture (AFF) is an atypical low-energy subtrochanteric and diaphyseal femoral fracture. Even if bone fusion is achieved in patients with AFF, the risk of AFF in the contralateral femur must be considered. This study aimed to investigate the factors affecting complete AFF in the contralateral femur and conservatively treated incomplete AFF., Subject and Methods: Radiographs of 111 femurs in 104 AFF cases were examined, and the femurs were classified as follows: 85 contralateral femurs with complete AFF; 18 contralateral femurs with incomplete AFF; 8 femurs with incomplete AFF without surgical treatment. Various patients' clinical data were collected, and we investigated the factors affecting the second complete AFF., Results: Complete fractures occurred in 10 (9.7%) of 103 femurs without incomplete AFF at the first visit and in 3 (37.5%) of 8 femurs with incomplete AFF. The Kaplan-Meier curve revealed that lateral cortical bone thickening and thigh pain were associated with significantly poorer prognoses (p = 0.026 and p = 0.013, respectively). Multivariate analyses revealed that eldecalcitol usage after AFF onset (p = 0.0094) and previous use of bisphosphonate or denosumab (p = 0.0126) were protective factors for second complete AFF and that the presence of thigh pain (p = 0.0134) was a risk factor for second complete AFF., Conclusions: Eldecalcitol administration after bone union of first AFF may prevent AFF recurrence. In addition, painful incomplete AFF has a high risk of developing a complete fracture., (© 2023 The Author(s). Published by S. Karger AG, Basel.)

To better understand the evolution of the SARS-CoV-2 Omicron subvariants, we performed molecular evolutionary analyses of the spike ( S ) protein gene/S protein using advanced bioinformatics technologies. First, time-scaled phylogenetic analysis estimated that a common ancestor of the Wuhan, Alpha, Beta, Delta variants, and Omicron variants/subvariants diverged in May 2020. After that, a common ancestor of the Omicron variant generated various Omicron subvariants over one year. Furthermore, a chimeric virus between the BM.1.1.1 and BJ.1 subvariants, known as XBB, diverged in July 2021, leading to the emergence of the prevalent subvariants XBB.1.5 and XBB.1.16. Next, similarity plot (SimPlot) data estimated that the recombination point (breakpoint) corresponded to nucleotide position 1373. As a result, XBB.1.5 subvariants had the 5' nucleotide side from the breakpoint as a strain with a BJ.1 sequence and the 3' nucleotide side as a strain with a BM.1.1.1 sequence. Genome network data showed that Omicron subvariants were genetically linked with the common ancestors of the Wuhan and Delta variants, resulting in many amino acid mutations. Selective pressure analysis estimated that the prevalent subvariants, XBB.1.5 and XBB.1.16, had specific amino acid mutations, such as V445P, G446S, N460K, and F486P, located in the RBD when compared with the BA.4 and BA.5 subvariants. Moreover, some representative immunogenicity-associated amino acid mutations, including L452R, F486V, R493Q, and V490S, were also found in these subvariants. These substitutions were involved in the conformational epitopes, implying that these mutations affect immunogenicity and vaccine evasion. Furthermore, these mutations were identified as positive selection sites. These results suggest that the S gene/S protein Omicron subvariants rapidly evolved, and mutations observed in the conformational epitopes may reduce the effectiveness of the current vaccine, including bivalent vaccines such as mRNA vaccines containing the BA.4/BA.5 subvariants.

Recently, there has been increasing interest in medial meniscal extrusion (MME), but few reports have evaluated MME via X-ray. In this study, the amount of MME and meniscal height at the medial border of the tibia were measured via X-ray with gradation processing. The extrusion length divided by the meniscal height yields the meniscal extrusion ratio, which was used as an index. In addition, the medial meniscal length of the part protruding from the medial border of the tibia on MRI was measured as an absolute value. Then, the correlation between the meniscal extrusion ratio and the amount of MME on MRI was examined, and there was a strong correlation between the meniscal extrusion ratio via X-ray and the amount of MME on MRI (correlation coefficient 0.860, p < 0.0001). The cut-off value of the meniscal extrusion ratio via X-ray for positive meniscal extrusion on MRI was 0.50, with an AUC of 0.9825, sensitivity of 0.9063, and specificity of 0.8663. From the present study, it was possible to measure the extrusion length and meniscal height via gradation processing, with X-ray and without MRI, and to calculate the meniscal extrusion ratio, which strongly correlates with the amount of MME on MRI.

Few evolutionary studies of the human respiratory virus (HRV) have been conducted, but most of them have focused on HRV3. In this study, the full-length fusion (F) genes in HRV1 strains collected from various countries were subjected to time-scaled phylogenetic, genome population size, and selective pressure analyses. Antigenicity analysis was performed on the F protein. The time-scaled phylogenetic tree using the Bayesian Markov Chain Monte Carlo method estimated that the common ancestor of the HRV1 F gene diverged in 1957 and eventually formed three lineages. Phylodynamic analyses showed that the genome population size of the F gene has doubled over approximately 80 years. Phylogenetic distances between the strains were short (< 0.02). No positive selection sites were detected for the F protein, whereas many negative selection sites were identified. Almost all conformational epitopes of the F protein, except one in each monomer, did not correspond to the neutralising antibody (NT-Ab) binding sites. These results suggest that the HRV1 F gene has constantly evolved over many years, infecting humans, while the gene may be relatively conserved. Mismatches between computationally predicted epitopes and NT-Ab binding sites may be partially responsible for HRV1 reinfection and other viruses such as HRV3 and respiratory syncytial virus., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023 The Author(s). Published by Elsevier B.V. All rights reserved.)

Age-related macular degeneration (AMD) is one of an increasing number of diseases that causes irreversible impairment and loss of vision in the elderly. AMD occurs by oxidative stress-mediated apoptosis of retinal pigment epithelium cells. The onset of AMD may be positively correlated with the exposure to blue light. We screened food-derived carotenoids for cytoprotective action against blue light irradiation using human ARPE-19 retinal pigment epithelium cells. This study revealed that blue light irradiation triggered apoptosis and oxidative stress in all- trans -retinal (atRAL)-exposed ARPE-19 cells by generating singlet oxygen ( 1 O 2 ), leading to significant cell death. We found that astaxanthin, a potent anti-oxidative xanthophyll abundant in several marine organisms including microalgae, salmon, and shrimp, significantly suppresses blue light-induced apoptotic cell death of atRAL-exposed ARPE-19 cells by scavenging 1 O 2 . Mechanistic studies using the blue-light irradiated cells also demonstrated that the cytoprotective effects of astaxanthin can be attributed to scavenging of 1 O 2 directly. Our results suggest the potential value of astaxanthin as a dietary strategy to prevent blue light-induced retinal degeneration including AMD.

The purpose of this study was to propose a novel classification of varus thrust based on gait analysis with inertial motion sensor units (IMUs) in patients with medial knee osteoarthritis (MKOA). We investigated thigh and shank acceleration using a nine-axis IMU in 69 knees with MKOA and 24 (control) knees. We classified varus thrust into four phenotypes according to the relative medial-lateral acceleration vector patterns of the thigh and shank segments: pattern A (thigh medial, shank medial), pattern B (medial, lateral), pattern C (lateral, medial), and pattern D (lateral, lateral). Quantitative varus thrust was calculated using an extended Kalman filter-based algorithm. We compared the differences between our proposed IMU classification and the Kellgren-Lawrence (KL) grades for quantitative varus thrust and visible varus thrust. Most of the varus thrust was not visually perceptible in early-stage OA. In advanced MKOA, increased proportions of patterns C and D with lateral thigh acceleration were observed. Quantitative varus thrust was significantly increased stepwise from patterns A to D. This novel IMU classification has better clinical utility due to its ability to detect subtle kinematic changes that cannot be captured with conventional motion analysis even in the early stage of MKOA.

Despite the increasing evidence of the clinical impact of Pseudomonas -derived cephalosporinase (PDC) sequence polymorphisms, the molecular evolution of its encoding gene, bla PDC , remains elusive. To elucidate this, we performed a comprehensive evolutionary analysis of bla PDC . A Bayesian Markov Chain Monte Carlo phylogenetic tree revealed that a common ancestor of bla PDC diverged approximately 4660 years ago, leading to the formation of eight clonal variants (clusters A-H). The phylogenetic distances within clusters A to G were short, whereas those within cluster H were relatively long. Two positive selection sites and many negative selection sites were estimated. Two PDC active sites overlapped with negative selection sites. In docking simulation models based on samples selected from clusters A and H, piperacillin was bound to the serine and the threonine residues of the PDC active sites, with the same binding mode for both models. These results suggest that, in P. aeruginosa , bla PDC is highly conserved, and PDC exhibits similar antibiotic resistance functionality regardless of its genotype.

Molecular interactions between respiratory syncytial virus (RSV) fusion protein (F protein) and the cellular receptor Toll-like receptor 4 (TLR4) and myeloid differentiation factor-2 (MD-2) protein complex are unknown. Thus, to reveal the detailed molecular interactions between them, in silico analyses were performed using various bioinformatics techniques. The present simulation data showed that the neutralizing antibody (NT-Ab) binding sites in both prefusion and postfusion proteins at sites II and IV were involved in the interactions between them and the TLR4 molecule. Moreover, the binding affinity between postfusion proteins and the TLR4/MD-2 complex was higher than that between prefusion proteins and the TLR4/MD-2 complex. This increased binding affinity due to conformational changes in the F protein may be able to form syncytium in RSV-infected cells. These results may contribute to better understand the infectivity and pathogenicity (syncytium formation) of RSV.

DNA gyrase plays important roles in genome replication in various bacteria, including Pseudomonas aeruginosa . The gyrA gene encodes the gyrase subunit A protein (GyrA). Mutations in GyrA are associated with resistance to quinolone-based antibiotics. We performed a detailed molecular evolutionary analyses of the gyrA gene and associated resistance to the quinolone drug, ciprofloxacin, using bioinformatics techniques. We produced an evolutionary phylogenetic tree using the Bayesian Markov Chain Monte Carlo (MCMC) method. This tree indicated that a common ancestor of the gene was present over 760 years ago, and the offspring formed multiple clusters. Quinolone drug-resistance-associated amino-acid substitutions in GyrA, including T83I and D87N, emerged after the drug was used clinically. These substitutions appeared to be positive selection sites. The molecular affinity between ciprofloxacin and the GyrA protein containing T83I and/or D87N decreased significantly compared to that between the drug and GyrA protein, with no substitutions. The rate of evolution of the gene before quinolone drugs were first used in the clinic, in 1962, was significantly lower than that after the drug was used. These results suggest that the gyrA gene evolved to permit the bacterium to overcome quinolone treatment.

Purpose: Patients undergoing knee surgery are at high risk for deep vein thrombosis (DVT), which is infrequent but potentially life-threatening. It has not been identified how to efficiently detect high-risk DVT while minimizing bleeding complications from anticoagulation. We hypothesized that the degree of activation of thrombotic markers may correlate with the size of the thrombus. Therefore, we investigated the correlation between thrombotic markers and DVT thrombus volume in patients after knee surgery., Methods: This retrospective study involved 29 patients who underwent around knee osteotomy or total / unicompartmental knee arthroplasty from 2018 to 2020. Fibrin monomer complex (FMC) at 1, and 7 days after surgery, and D-dimer at 4, and 7 days after surgery were investigated. In addition, the volume of DVT was estimated with ultrasonography at the 7 days after surgery. Body mass index, surgical time, and total volume of blood loss were also evaluated. Factors related to thrombus volume were examined statistically., Results: Nine patients (31.0%) exhibited asymptomatic distal DVT, whereas 1 patient (3.4%) experienced asymptomatic proximal DVT. No patients had pulmonary embolism. Statistical analysis showed that only FMC concentration on postoperative day 1 was significantly correlated with thrombus volume (p <  0.001, 95% confidence interval 0.41 to 0.839, r = 0.679)., Conclusions: The FMC concentration was a useful early indicator of deep vein thrombosis after knee surgery. Monitoring the FMC concentration could enable selective identification of patients with a high thrombus volume, which is associated with a high risk for pulmonary embolism., (© 2022. The Author(s).)

Polyphenols, especially catechol-type polyphenols, exhibit lysyl oxidase-like activity and mediate oxidative deamination of lysine residues in proteins. Previous studies have shown that polyphenol-mediated oxidative deamination of lysine residues can be associated with altered electrical properties of proteins and increased crossreactivity with natural immunoglobulin M antibodies. This interaction suggested that oxidized proteins could act as innate antigens and elicit an innate immune response. However, the structural basis for oxidatively deaminated lysine residues remains unclear. In the present study, to establish the chemistry of lysine oxidation, we characterized oxidation products obtained via incubation of the lysine analog N-biotinyl-5-aminopentylamine with eggshell membranes containing lysyl oxidase and identified a unique six-membered ring 2-piperidinol derivative equilibrated with a ring-open product (aldehyde) as the major product. By monitoring these aldehyde-2-piperidinol products, we evaluated the lysyl oxidase-like activity of polyphenols. We also observed that this reaction was mediated by some polyphenols, especially o-diphenolic-type polyphenols, in the presence of copper ions. Interestingly, the natural immunoglobulin M monoclonal antibody recognized these aldehyde-2-piperidinol products as an innate epitope. These findings establish the existence of a dynamic equilibrium of oxidized lysine and provide important insights into the chemopreventive function of dietary polyphenols for chronic diseases., Competing Interests: Conflict of interest The authors declare that they have no conflicts of interest with the contents of this article., (Copyright © 2021 The Authors. Published by Elsevier Inc. All rights reserved.)

Sulfites are commonly used as a preservative and antioxidant additives in the food industry. Sulfites are absorbed by the gastrointestinal tract and distributed essentially to all body tissues. Although sulfites have been believed to be safe food additives, some studies have shown that they exhibit adverse effects in various tissues. In this study, we examined the cytotoxic effect of sodium sulfite (Na 2 SO 3 ) against rat gastric mucosal cells (RGM1) and further investigated its underlying molecular mechanism. We demonstrated that exposure to Na 2 SO 3 exerts significant cytotoxicity in RGM1 cells through induction of oxidative stress. Exposure of RGM1 cells to Na 2 SO 3 caused a significant formation of protein carbonyls and 8-hydroxy-2'-deoxyguanosine, major oxidative stress markers, with a concomitant accumulation of carbonylated protein-related aggregates. Furthermore, we found that incubation of lysozyme with Na 2 SO 3 evokes protein carbonylation and aggregation via the metal ion-catalyzed free radical formation derived from Na 2 SO 3 . Our results suggest that Na 2 SO 3 might lead to gastric tissue injury via induction of oxidative stress by the formation of Na 2 SO 3 -related free radicals.

Proteins are oxidatively modified by a large number of reactive species including reactive oxygen species, lipid peroxidation-derived aldehydes, and reducing sugars. Among divergent oxidative modifications, the introduction of carbonyl groups such as aldehyde, ketone, and lactam into the amino acid side chains of proteins is a major hallmark for oxidative damage to proteins, and is termed "protein carbonylation". Detection and quantification of protein carbonyls are commonly performed to determine the level of oxidative stress in the context of cellular damage, aging, and several age-related disorders. This review focuses on the molecular mechanisms and biological implications of protein carbonylation, and also presents current analytical approaches for determining and characterizing carbonylated proteins.

Objectives: The purpose of this study was to clarify the diagnostic accuracy of the mobile assessment of varus thrust using inertial measurement units (IMUs)., Methods: A total of 80 knees in 49 patients were enrolled in this study. On visual analysis of gait to determine the presence or absence of varus thrust, 23 knees were assigned to the Present group, 17 to the Ambiguous group, and 40 to the Absent group. The peak knee varus angular velocities (PVVs), measured by quantitative gait analysis using nine-axis IMUs, were compared between these three groups. A receiver operating characteristic curve for the relationship between the visual assessment of varus thrust (Present and Ambiguous) and the measured PVV was created, and the cut-off PVV for visualized varus thrust was determined as the highest point for both sensitivity and specificity., Results: The mean PVVs were significantly different between the three groups (Present, 47.7 ± 8.2 degree/s, Ambiguous, 34.1 ± 10.5 degree/s, and Absent, 28.1 ± 8.3 degree/s, respectively, ANOVA P=0.000). The PVV cut-off value for visualized varus thrust was 28.1 degree/s, yielding a sensitivity of 0.957 and a specificity of 0.579., Conclusions: A PVV <28.1 degree/s is useful for ruling out varus thrust during gait. This quantitative varus thrust assessment method using IMUs has clinical utility as a screening test., Competing Interests: CONFLICTS OF INTEREST: This research did not receive any specific funding from the public, commercial, or not-for-profit sectors., (©2021 The Japanese Association of Rehabilitation Medicine.)

Purpose: Teriparatide is sometimes used in the treatment of atypical femoral fracture (AFF). Even if bone union is achieved, orthopedic physicians must consider the risk of relapse. This study aimed to investigate the factors affecting AFF recurrence, and to determine the appropriate treatment for osteoporosis after bone union., Methods: One hundred thirty-one consecutive AFFs in 113 Japanese patients were included. Eleven patients had AFF in the unaffected limb (9 patients) after the first AFF or re-fracture at the original fracture site (2 patients) after bone union of the first AFF was confirmed. We divided all patients into two groups: the second fracture group (22 AFFs in 11 patients) and non-second fracture group (109 AFFs in 102 patients). We compared clinical information between the 2 groups and investigated the factors affecting AFF recurrence using the Student t-, Welch t-, and chi-square tests., Results: Although there was no significant difference in clinical characteristics between the 2 groups, multivariate analysis of factors associated with AFF recurrence identified short duration of treatment with teriparatide and active vitamin D 3 (p = 0.0408 and 0.0366, respectively) as risk factors. Even in the analysis excluding subtrochanteric AFF, short periods of teriparatide and active vitamin D 3 administration were observed as risk factors (p = 0.0484 and 0.0346, respectively)., Conclusion: The administration of teriparatide for as long as possible after occurrence first AFF and the use of active vitamin D 3 after completion of teriparatide therapy may be the most effective strategy to prevent the recurrence of AFF., (Copyright © 2020 Elsevier Inc. All rights reserved.)

Objectives: Differences in mechanisms of subtrochanteric and diaphyseal atypical femoral fractures (AFFs) are speculated in studies that analyzed differences in the patients' background. However, the etiologies of each type of AFF have not been studied in detail. This study aimed to investigate the nature and etiologies of the risk factors for diaphyseal AFFs., Materials and Methods: Eighty consecutive Japanese patients with 91 diaphyseal AFFs (AFF group) and 110 age-matched women with osteoporosis (non-AFF control group) were included. Their clinical data were compared; factors affecting AFFs were investigated, and the etiologies of the risk factors for diaphyseal AFFs were examined., Results: Multivariate analysis revealed that femoral serrated changes, bisphosphonate or denosumab usage, and lateral and anterior femoral curvatures were risk factors for diaphyseal AFFs (p < 0.0011, p = 0.0137, and p < 0.0001, respectively). Multivariate analyses revealed that serrated changes and low serum 25(OH)D levels affected the lateral curvature (p = 0.0088 and 0.0205, respectively), while serrated changes affected the anterior curvature (p = 0.0006), each significantly affected the femoral curvature. High serum calcium (Ca) levels, lateral femoral curvature, and anterior femoral curvature were predictors of serrated changes (p = 0.0146, 0.0002, and 0.0098, respectively)., Conclusion: Risk factors for diaphyseal AFFs were bone resorption inhibitor usage, a strong femoral curvature, and serrated changes. Low serum 25(OH)D levels and serrated changes are risk factors for lateral curvature, while a high serum Ca level is a risk factor for serrated changes., (© 2021 The Author(s) Published by S. Karger AG, Basel.)

Background: Spontaneous osteonecrosis of the knee (SONK) is one of the acute knee pain disorders arising in elderly patients. The presence of knee varus alignment and the size of necrotic area have been reported as the negative prognostic factors in prior studies. However, no previous study has yet clarified the radiological analysis of the lower extremity in SONK compared with that in osteoarthritis. The purpose of this study was therefore to identify the radiographic findings of the lower extremity in SONK., Methods: Sixty-three knees of Kellgren-Lawrence classification grade 1 or 2 without any trauma treated between April 2012 and March 2014 were enrolled in this study. These knees were divided into two groups according to their magnetic resonance imaging (MRI) findings: SONK group (31 knees) and OA group (32 knees). Using a long leg standing X-ray, femorotibial angle (FTA), mechanical axis deviation (MAD), mechanical lateral distal femoral angle (mLDFA), medial proximal tibial angle (MPTA) and joint line convergent angle (JLCA) were compared between groups. Correlation between each parameter and the width ratio (WR) of the necrotic lesion were analyzed., Results: FTA, MAD, MPTA and JLCA showed significant differences between the SONK and OA groups. In the SONK group, FTA was positively correlated with WR, and, MAD and MPTA was negatively correlated with WR., Conclusions: Compared with OA, SONK is associated with a significantly larger varus deformity at the proximal tibia, and larger joint play in the coronal plane., Competing Interests: Declaration of competing interest This research did not receive any specific grant from funding agencies in the public, commercial, or not-for-profit sectors. The authors declare no competing interests in relation to this study., (Copyright © 2020 Elsevier B.V. All rights reserved.)

The purpose of this study was to assess the correlation between tissue volume and blood flow of the flap in an animal model. Using animal model, tissue volume can be attenuated, and precise change of blood flow could be evaluated. We further investigate the relationship between blood flow and vascular density in the tissue. In this study, we assessed flap conductance (ml/min/mm Hg) as to evaluate the conductivity of blood flow into the flap. Japanese white rabbit was used (n = 7) for this study. The amount of blood flow of jejunal and latissimus dorsi muscle (LD) flaps was measured while removing the distal portion of the flap sequentially. Conductance at each time was calculated from blood pressure and blood flow volume. The tissue volume at each time was also measured. The correlation between conductance and volume was analyzed using a linear mixed model. Immunohistochemical evaluation of microvessel densities (MVD) in these tissues was also performed for CD31/PECAM1 positive area. Conductance and tissue volume were significantly correlated in both jejunal and LD flaps. As the volume increases by 1 cm 3 , the conductance increased significantly by 0.012 ml/min/mm Hg in jejunum, and by 0.0047 ml/min/mm Hg in LD. Mean MVD was 1.15 ± 0.52% in the jejunum and 0.37 ± 0.29% in the LD muscle. In this study, we revealed that flap conductance is proportional to volume and proportional constant is different between the type of tissue. It suggests that the difference of MVD creates the unique conductance of each tissue., Competing Interests: The authors have no conflicts of interest, whether they are financial or related to any other relationships, to disclose.

Introduction: Postmenopausal osteoporosis and dyslipidemia are well-known skeletal and metabolic changes in middle-aged women. We investigated the effects of combined treatments with a selective estrogen receptor modulator (SERM) and exercise on bone and fat parameters in ovariectomized (OVX) rats., Materials and Methods: Sixteen-week-old female Sprague-Dawley rats underwent bilateral ovariectomy, and rats were randomized to BZA (bazedoxifene at 0.3 mg/kg/day), Exe (treadmill exercise at 12-15 m/min, 60 min/day, 5 days/week), Comb (BZA and Exe), and Cont (control treated with vehicle and no exercise) groups 8 weeks after ovariectomy. After 4 or 8 weeks of treatment, bone mineral density (BMD) of the total femur and lumbar spine and whole-body percentage fat mass were determined by dual-energy X-ray absorptiometry, and mechanical testing of the femoral shaft, and bone and fat histomorphometric analyses of the proximal tibia were performed., Results: Treadmill exercise had decreased bone marrow adipocytes from 4 weeks of treatment and whole-body percentage fat mass at 8 weeks. BZA increased BMD at the lumbar spine and decreased the whole-body percentage fat mass from 4 weeks and bone marrow adipocytes at 8 weeks. Combination therapy increased BMD for the lumbar spine and decreased bone marrow adipocytes and whole-body percentage fat mass from 4 weeks., Conclusion: Combination therapy with BZA and exercise appears effective to improve bone and fat parameters in OVX rats.

Scope: Nonalcoholic fatty liver disease (NAFLD) has emerged as the most common chronic liver disease worldwide, defined by hepatic over-accumulation of lipids without significant ethanol consumption. Pharmacological or bioactive food ingredients that suppress hepatic lipid accumulation through AMP-activated protein kinase (AMPK) signaling, which plays a critical role in the regulation of lipid metabolism, are searched., Methods and Results: It is found that tomatidine, the aglycone of α-tomatine abundant in green tomatoes, significantly inhibits palmitate-provoked lipid accumulation and stimulates phosphorylation of AMPK and acetyl-CoA carboxylase 1 (ACC1) in human HepG2 hepatocytes. The results also indicate that tomatidine can enhance triglyceride turnover and decline in lipogenesis by upregulating adipose triglyceride lipase (ATGL) and downregulating fatty acid synthase (FAS) via the AMPK signaling-dependent regulation of transcription factors, element-binding protein-1c (SREBP-1c) and forkhead box protein O1 (FoxO1). Furthermore, mechanistic studies demonstrate that tomatidine-stimulated AMPK phosphorylation is due to CaMKKβ activation in response to an increase in intracellular Ca 2+ concentration. Finally, it is discovered that tomatidine functions as an agonist for vitamin D receptor to elicit AMPK-dependent suppression of lipid accumulation., Conclusion: The in vitro study suggests the potential efficacy of tomatidine as a preventive and therapeutic treatment in obesity-related fatty liver diseases., (© 2019 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.)

Short-chain fatty acids (SCFAs) such as acetate, propionate, and butyrate are generated by gut microbial fermentation of dietary fiber. SCFAs may exert multiple beneficial effects on human lipid and glucose metabolism. However, their actions and underlying mechanisms are not fully elucidated. In this study, we examined the direct effects of propionate on hepatic glucose and lipid metabolism using human HepG2 hepatocytes. Here, we demonstrate that propionate at a physiologically-relevant concentration effectively suppresses palmitate-enhanced glucose production in HepG2 cells but does not affect intracellular neutral lipid levels. Our results indicated that propionate can decline in gluconeogenesis by down-regulation of glucose-6-phosphatase (G6Pase) and phosphoenolpyruvate carboxykinase (PEPCK) through activation of AMP-activated protein kinase (AMPK), which is a major regulator of the hepatic glucose metabolism. Mechanistic studies also revealed that propionate-stimulated AMPK phosphorylation can be ascribed to Ca 2+ /calmodulin-dependent protein kinase kinase β (CaMKKβ) activation in response to an increase in intracellular Ca 2+ concentration. Moreover, siRNA-mediated knockdown of the propionate receptor GPR43 prevented propionate-inducible activation of AMPK and abrogates the gluconeogenesis-inhibitory action. Thus, our data indicate that the binding of propionate to hepatic GPR43 elicits CaMKKβ-dependent activation of AMPK through intracellular Ca 2+ increase, leading to suppression of gluconeogenesis. The present study suggests the potential efficacy of propionate in preventive and therapeutic management of diabetes., (Copyright © 2019 Elsevier Inc. All rights reserved.)

The interactions between neuronal, glial, and vascular cells play a key role in regulating blood flow in the retina. In the present study, we examined the role of the interactions between neuronal and glial cells in regulating the retinal vascular tone in rats upon stimulation of retinal neuronal cells by intravitreal injection of N-methyl-d-aspartic acid (NMDA). The retinal vascular response was assessed by measuring the diameter of the retinal arterioles in the in vivo fundus images. Intravitreal injection of NMDA produced retinal vasodilation that was significantly diminished following the pharmacological inhibition of nitric oxide (NO) synthase (nNOS), loss of inner retinal neurons, or intravitreal injection of glial toxins. Immunohistochemistry revealed the expression of nNOS in ganglion and calretinin-positive amacrine cells. Moreover, glial toxins significantly prevented the retinal vasodilator response induced by intravitreal injection of NOR3, an NO donor. Mechanistic analysis revealed that NO enhanced the production of vasodilatory prostanoids and epoxyeicosatrienoic acids in glial cells in a ryanodine receptor type 1-dependent manner, subsequently inducing the retinal vasodilator response. These results suggest that the NO released from stimulated neuronal cells acts as a key messenger in neuron-glia signaling, thereby causing neuronal activity-dependent and glial cell-mediated vasodilation in the retina., Competing Interests: The authors report no conflicts of interest. The funders had no role in the design of the study; in the collection, analyses, or interpretation of data; in the writing of the manuscript, or in the decision to publish the results.

We investigated the effects of ibandronate, a bisphosphonate; eldecalcitol, an active vitamin D3 analogue; and combination treatment with both agents on secondary osteoporosis and arthritis using rats with adjuvant-induced arthritis. Arthritis was induced in 8-week-old male Lewis rats. Rats were randomized into four treatment groups and an untreated normal control group: ibandronate, eldecalcitol, ibandronate + eldecalcitol, vehicle, and control. Paw thickness was measured to evaluate arthritis. Joint destruction was evaluated histomorphometrically by the ankle joint stained with Fast Green and safranin O. The femur and lumbar spine were scanned using dual-energy X-ray absorptiometry, and the distal femur was scanned using micro-computed tomography for bone mineral density (BMD) and trabecular microstructural evaluations. Ibandronate and/or eldecalcitol increased BMD in both the lumbar vertebrae and femur and improved several microstructural parameters (bone volume/total volume, structure model index, trabecular number, and trabecular separation of the distal femur). In addition, there was an additive effect of combination treatment compared with single treatments for most trabecular parameters, including BMD and bone volume. However, ibandronate and/or eldecalcitol did not inhibit arthritis and joint destruction. Combination treatment with ibandronate and eldecalcitol may be effective for secondary osteoporosis associated with arthritis.

Objectives: Rheumatoid arthritis (RA) is characterized by chronic inflammation of the synovium, progressive erosion of the articular cartilage, and joint destruction. RA also causes secondary osteoporosis and muscle wasting. We investigated the effects of ibandronate (IBN), a bisphosphonate; eldecalcitol (ELD), an active vitamin D3 derivative; and combination treatment with both agents on secondary osteoporosis and muscle wasting using adjuvant-induced arthritis rats., Methods: Arthritis was induced in 8-week-old male Lewis rats. Rats were randomized into 4 treatment groups and an untreated normal control group: IBN (subcutaneously, once every 2 weeks, 10 μg/kg), ELD (orally, once daily, 30 ng/kg/day), IBN + ELD, vehicle, and control. Paw thickness measurements were performed for evaluation of arthritis. The femur was scanned using dual-energy X-ray absorptiometry. Cross-sectional areas of left tibialis and anterior muscle fibers and the expression of MuRF1, atrogin-1, MyoD, and myogenin in the gastrocnemius muscle were measured to evaluate muscle wasting., Results: IBN and/or ELD increased bone mineral density (BMD) in the femur. In addition, there was an additive effect of combination treatment compared with single treatments for BMD. However, IBN and/or ELD did not inhibit muscle wasting in adjuvant-induced arthritis rats., Conclusions: Combination treatment with IBN and ELD may be effective for secondary osteoporosis associated with RA. Other treatments are necessary for muscle wasting associated with RA. Studies in humans are needed to confirm these findings.

Obesity, a principal risk factor for the development of diabetes mellitus, heart disease, and hypertension, is a growing and serious health problem all over the world. Leptin is a weight-reducing hormone produced by adipose tissue, which decreases food intake via hypothalamic leptin receptors (Ob-Rb) and the Janus kinase 2/signal transducer and activator of transcription 3 (JAK2/STAT3) signaling pathway. Protein tyrosine phosphatase 1B (PTP1B) negatively regulates leptin signaling by dephosphorylating JAK2, and the increased activity of PTP1B is implicated in the pathogenesis of obesity. Hence, inhibition of PTP1B may help prevent and reduce obesity. In this study, we revealed that phenethyl isothiocyanate (PEITC), a naturally occurring isothiocyanate in certain cruciferous vegetables, potently inhibits recombinant PTP1B by binding to the reactive cysteinyl thiol. Moreover, we found that PEITC causes the ligand-independent phosphorylation of Ob-Rb, JAK2, and STAT3 by inhibiting cellular PTP1B in differentiated human SH-SY5Y neuronal cells. PEITC treatment also induced nuclear accumulation of phosphorylated STAT3, resulting in enhanced anorexigenic POMC expression and suppressed orexigenic NPY/AGRP expression. We demonstrated that oral administration of PEITC to mice significantly reduces food intake, and stimulates hypothalamic leptin signaling. Our results suggest that PEITC might help prevent and improve obesity., Competing Interests: The authors have declared that no competing interests exist.

Diabetes mellitus causes secondary osteoporosis and muscle atrophy. The ability of alfacalcidol (ALF) and exercise (Exe) to inhibit osteoporosis and muscle atrophy in type 2 diabetes mellitus (T2DM) model rats was examined. Twenty-week-old Otsuka Long-Evans Tokushima Fatty rats were randomized to ALF (orally 0.1 μg/kg/day), Exe (treadmill exercise at 10 m/min, 60 min/day, 5 days/week), Comb (ALF and Exe), and Cont (T2DM control treated with vehicle and no exercise) groups (n = 8-10 per group). Sedentary Long-Evans Tokushima Otsuka rats were used as a non-hyperphagic control. After treatment for 2 or 6 weeks, blood glucose (BG) levels, cross-sectional area (CSA) of tibialis anterior muscle fibers, femoral bone mineral density (BMD), and relative quantities of muscle anabolic markers (Pax7, MyoD, and myogenin) and catabolic markers (Atrogin-1, MuRF1, and REDD1) of the soleus muscle assessed by real-time polymerase chain reaction assays were measured. Exe and Comb treatments for 6 weeks decreased BG levels compared with those of the Cont group. ALF, Exe, and Comb treatments for 2 and 6 weeks recovered the CSA compared with that of the Cont group. ALF and Comb treatments for 6 weeks increased femoral BMDs compared with those of the Cont group. After 2 weeks of treatment, Comb treatment increased MyoD expression and decreased MuRF1 expression. ALF or Exe monotherapy significantly decreased Atrogin-1 or MuRF1 expression after 2 weeks of treatment, respectively. After 6 weeks of treatment, ALF and Comb treatments decreased Atrogin-1 and REDD1. These results demonstrate that a combination of ALF and Exe improved CSA from the early phase of treatment by stimulating skeletal muscle differentiation and suppressing muscle catabolic genes. Improvements in BG, BMD, and CSA were observed as long-term effects of the combination therapy. Continued suppression of muscle catabolic genes was observed as a background to these effects., Competing Interests: The authors have declared that no competing interests exist.

Scope: The prevalence of type 2 diabetes mellitus (T2DM) is increasing yearly worldwide. Glycemic control is the basis for the treatment of T2DM, as it can prevent the progress of associated complications. Spices possess various health beneficial effects on humans. The aim of this study is to search for spices that can promote glucose uptake and to elucidate the underlying molecular mechanism(s)., Methods and Results: Among 24 spice extracts, the extracts from black pepper and white pepper significantly increase glucose uptake in L6 myotubes. Piperine is found to be the active compound in these extracts. Treatment of myotubes with piperine induces the translocation of glucose transporter 4 (GLUT4) to the plasma membrane by phosphorylation of AMP-activated protein kinase (AMPK). Piperine increases the intracellular Ca 2+ level and reactive oxygen species (ROS) generation through transient receptor potential vanilloid channel 1 (TRPV1), followed by activation of Ca 2+ /calmodulin-dependent protein kinase kinase-beta (CaMKKβ) as the upstream events for AMPK phosphorylation. Furthermore, oral administration of piperine to Wistar rats at 0.01 and 0.1 mg kg -1 body weight decreases postprandial hyperglycemia accompanied by GLUT4 translocation and AMPK phosphorylation., Conclusion: Piperine in pepper prevents hyperglycemia by GLUT4 translocation through CaMKKβ/AMPK signaling via TRPV1-dependent increase in the intracellular Ca 2+ level and ROS generation., (© 2018 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.)

We evaluated the analgesic effects of minodronate, alendronate and pregabalin on mechanical and thermal allodynia, as well as changes in bone mineral density and skeletal muscle volume caused by chronic constriction injury (CCI) in an ovariectomized rat. Ovariectomy was performed on four-week-old female Wistar rats. Thereafter, at 8-weeks of age, the left sciatic nerve was ligated to create the chronic pain model (CCI limb), and sham surgery was performed on the right hindlimb. In all rats, either minodronate (0.15 mg/kg/week), alendronate (0.15 mg/kg/week), pregabalin (10 mg/kg/week), or their vehicle was administered for 2 weeks starting on the 0th day of CCI. Behavioral evaluations, with von Frey testing and the hot plate test, were performed on days 0, 7 and 14. After 2 weeks, bilateral femurs and tibialis anterior muscles were harvested for bone mineral density and cross sectional area measurements, respectively. Two weeks treatment with minodronate significantly improved mechanical and thermal allodynia evaluated by the von Frey and hot plate tests in the CCI limb (P < 0.05). Minodronate and alendronate treatment for 2 weeks significantly increased total femoral bone mineral density in the CCI limb compared with pregabalin or vehicle treatment (P < 0.01). Cross sectional area of the CCI limb in the minodronate group was significantly larger than that of the alendronate group (P < 0.05) and pregabalin group (P < 0.05). Two-week treatment with minodronate, but not alendronate or pregabalin, improved mechanical and thermal allodynia caused by CCI in ovariectomized rats.

Methylglyoxal, a highly reactive dicarbonyl compound, is formed as a by-product of glycolysis and plays an important role in the pathogenesis of diabetic complications, including diabetic retinopathy. However, it remains to be determined how methylglyoxal affects the regulatory mechanisms of retinal blood flow. In this study, we examined the effects of methylglyoxal on β 2 -adrenoceptor-mediated vasodilatory mechanisms in rat retinal arterioles. The retinal vasodilator responses were assessed by measuring the diameter of retinal arterioles in the fundus images. Intravitreal injection of methylglyoxal significantly diminished the vasodilation of retinal arterioles induced by the β 2 -adrenoceptor agonist salbutamol. The vasodilator effect of BMS-191011, a large-conductance Ca 2+ -activated K + (BK Ca ) channel opener, on retinal arterioles was also attenuated by methylglyoxal. In contrast, methylglyoxal had no significant effect on retinal vasodilator response to forskolin. Methylglyoxal attenuated retinal vasodilator response to salbutamol under blockade of BK Ca channels with iberiotoxin, an inhibitor of the channels. These results suggest that methylglyoxal attenuates β 2 -adrenoceptor-mediated retinal vasodilation by impairing the coupling of the β 2 -adrenoceptor to the guanine nucleotide-binding protein (Gs protein) and the function of the BK Ca channel. Increased methylglyoxal in the eyes may contribute to the impairment of regulatory mechanisms of retinal blood flow in patients with diabetic retinopathy.

Pyrroloquinoline quinone (PQQ), a redox-active o-quinone found in various foods and mammalian tissues, has received an increasing amount of attention because of a number of health benefits that can be attributed to its ability to enhance mitochondrial biogenesis. However, its underlying molecular mechanism remains incompletely understood. We have now established that the exposure of mouse NIH/3T3 fibroblasts to a physiologically relevant concentration of PQQ significantly stimulates mitochondrial biogenesis. The exposure of NIH/3T3 cells to 10-100 nM PQQ for 48 h resulted in increased levels of Mitotracker staining, mitochondrial DNA content, and mitochondrially encoded cytochrome c oxidase subunit 1 (MTCO1) protein. Moreover, we observed that PQQ treatment induces deacetylation of the peroxisome proliferator-activated receptor-γ-coactivator 1α (PGC-1α) and facilitates its nuclear translocation and target gene expression but does not affect its protein levels, implying increased activity of the NAD + -dependent protein deacetylase sirtuin 1 (SIRT1). Indeed, treatment with a SIRT1 selective inhibitor, EX-527, hampered the ability of PQQ to stimulate PGC-1α-mediated mitochondrial biogenesis. We also found that the PQQ treatment caused a concentration-dependent increase in the cellular NAD + levels, but not the total NAD + and NADH levels. Our results suggest that PQQ-inducible mitochondrial biogenesis can be attributed to activation of the SIRT1/PGC-1α signaling pathway by enhancing cellular NAD + formation.

Objectives: Glucocorticoid (GC) treatment inhibits activation of runt-related transcription factor 2 (Runx2), which is essential for osteoblast differentiation from stem cells. As a result, GC treatment results in bone loss, GC-induced osteoporosis (GIO), elevated fracture risk, and delayed bone healing. Bisphosphonates such as alendronate (ALN) are recommended for treating or preventing GIO, and low-intensity pulsed ultrasound (LIPUS) facilitates fracture healing and maturation of regenerated bone. Combined therapy with ALN and LIPUS may stimulate cancellous bone healing in GIO rats. Here, we examined the effect of ALN and LIPUS on cancellous bone osteotomy repair in the proximal tibia of GIO rats., Methods: Prednisolone (10 mg/kg body weight/day) was administered for 4 weeks to induce GIO in 6-month-old female Sprague-Dawley rats. Tibial osteotomy was then performed and daily subcutaneous injection of ALN (1-μg/kg body weight) was subsequently administered alone or in combination with LIPUS (20 min/day) for 2 or 4 weeks., Results: ALN significantly increased bone mineral density (BMD) at 2 and 4 weeks, and ALN + LIPUS significantly increased BMD at 4 weeks. Bone union rates were significantly increased after 2 and 4 weeks ALN and ALN + LIPUS treatment. Lastly, ALN and ALN + LIPUS significantly increased the proportion of Runx2 positive cells at 4 weeks., Conclusions: ALN monotherapy and combined ALN and LUPUS treatment augmented BMD and stimulated cancellous bone repair with increased Runx2 expression at the osteotomy site in GIO rats. However, the combined treatment had no additional effect on cancellous bone healing compared to ALN monotherapy.

Polyphenols have received a significant amount of attention in disease prevention because of their unique chemical and biological properties. However, the underlying molecular mechanism for their beneficial effects remains unclear. We have now identified a polyphenol as a source of innate epitopes detected in natural IgM and established a unique gain-of-function mechanism in the formation of innate epitopes by polyphenol via the polymerization of proteins. Upon incubation with bovine serum albumin (BSA) under physiological conditions, several polyphenols converted the protein into the innate epitopes recognized by the IgM Abs. Interestingly, piceatannol, a naturally occurring hydroxylated analogue of a red wine polyphenol, resveratrol, mediated the modification of BSA, whose polymerized form was specifically recognized by the IgMs. The piceatannol-mediated polymerization of the protein was associated with the formation of a lysine-derived cross-link, dehydrolysinonorleucine. In addition, an oxidatively deaminated product, α-aminoadipic semialdehyde, was detected as a potential precursor for the cross-link in the piceatannol-treated BSA, suggesting that the polymerization of the protein might be mediated by the oxidation of a lysine residue by piceatannol followed by a Schiff base reaction with the ε-amino group of an unoxidized lysine residue. The results of this study established a novel mechanism for the formation of innate epitopes by small dietary molecules and support the notion that many of the beneficial effects of polyphenols could be attributed, at least in part, to their lysyl oxidase-like activity. They also suggest that resveratrol may have beneficial effects on human health because of its conversion to piceatannol.