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Purpose: To examine the hypothesis that an increase in response of postmenopausal bone to interval walking training (IWT) depends on baseline bone mineral densities (BMDs). Methods: Two hundred and thirty-four postmenopausal women (64±5 (SD) yr) with no medication for osteoporosis performed 5-month IWT, repeating fast and slow walking at ≥70% and ~40% peak aerobic capacity, respectively, for 3 minutes each per set, ≥5 sets/day, ≥4 days/week. They were recruited from those who had performed IWT ≥6 months before participating in the study so that their physical fitness and lifestyle-related disease symptoms had almost reached a steady state at the time of their participation. We measured BMDs for the lumbar spine (LS), bilateral femoral neck (FN), and bilateral total hip (TH) by dual-energy X-ray absorptiometry (DXA) before and after the intervention. We used a multiple regression analysis to identify significant independent factors for increasing BMDs after the intervention as baseline physical characteristics, exercise intensity, and exercise time during IWT were the candidates. For any bone site where the independent factor identified was singular, we analyzed the relationship between the identified factor vs increases in BMD after the intervention (ΔBMD) by the locally weighted scatterplot smoothing (LOWESS) method. Results: Almost all subjects completed the designated protocol with minimal adverse events. We found that significant determinants for increasing BMDs were the baseline BMDs for all bone sites, as well as age and body mass index for TH (all, P<0.02). Furthermore, the LOWESS trendline between ΔBMDs vs the baseline BMDs, divided equally into 10 bins for LS and FN, respectively, showed that ΔBMD responses (Y) were attenuated as the baseline BMDs (X) increased in the lower 4 bins and then showed a flat line (Y = ~0) in the remaining higher 6 bins for LS and FN. When the lower 4 bins and the higher 6 bins of the baseline BMD were pooled, respectively, BMDs significantly increased by 1.8% and 1.0% in the lower groups for LS and FN, respectively (both, P<0.001) while not in the higher groups after the intervention (both, P>0.3). Conclusions: IWT may be of benefit with minimal adverse events to postmenopausal women, although the effects were greater in those with lower baseline BMDs. Trial registration: UMIN000047428. https://rctportal.niph.go.jp/s/detail/um?trial_id=UMIN000047428#. [ABSTRACT FROM AUTHOR]

The aim of the present study was to investigate the perioperative and postoperative incidence of deep vein thrombosis (DVT) and validate the effectiveness of our own preventive treatment protocol for venous thromboembolism (VTE) occurrence in lower extremity arthroplasty patients. The subjects were 1,054 patients (mean age: 74.3 years) who underwent total hip arthroplasty (THA) or total knee arthroplasty (TKA) at our institutions between April 2014 and March 2017. We examined the frequencies of pre- and post-operative DVT by lower extremity Doppler images, and the incidence rate at proximal or distal regions as well as that according to preoperative DVT status were evaluated. Preoperative DVT was detected in 6.5% (69 cases) of our cohort and those were located 1.4% (15 cases) at proximal and 5.1% (54 cases) at distal regions. A significantly higher rate of postoperative DVT development was observed in preoperative DVT+ THA patients (P = 0.0075), but not in TKA patients only with a higher tendency (P = 0.56). The overall incidence of DVT up to 2 weeks after surgeries was 27.3% (288 cases); however, the rate in proximal femur regions was suppressed to 2.8% (30 cases), and there was no symptomatic pulmonary thromboembolism (PTE) case. The results demonstrated the importance of regular Doppler examination for early detection of postoperative DVT occurrence and the following immediate treatment initiation. Our own VTE preventive treatment protocol could reduce the development of proximal DVT, and the periodic monitoring as well as prompt treatment might prevent the fatal PTE. osteoarthritis (OA), rheumatoid arthritis (RA). [ABSTRACT FROM AUTHOR]

Sickle Cell Disease and ß-thalassemia, which are caused by defective or deficient adult ß-globin (HBB) respectively, are the most common serious genetic blood diseases in the world. Persistent expression of the fetal ß-like globin, also known as γ-globin, can ameliorate both disorders by serving in place of the adult ß-globin as a part of the fetal hemoglobin tetramer (HbF). Here we use CRISPR-Cas9 gene editing to explore a potential γ-globin silencer region upstream of the δ-globin gene identified by comparison of naturally-occurring deletion mutations associated with up-regulated γ-globin. We find that deletion of a 1.7 kb consensus element or select 350 bp sub-regions from bulk populations of cells increases levels of HbF. Screening of individual sgRNAs in one sub-region revealed three single guides that caused increases in γ-globin expression. Deletion of the 1.7 kb region in HUDEP-2 clonal sublines, and in colonies derived from CD34+ hematopoietic stem/progenitor cells (HSPCs), does not cause significant up-regulation of γ-globin. These data suggest that the 1.7 kb region is not an autonomous γ-globin silencer, and thus by itself is not a suitable therapeutic target for gene editing treatment of ß-hemoglobinopathies. [ABSTRACT FROM AUTHOR]

The Janus kinases (Jaks) are hubs in the signaling process of more than 50 cytokine or hormone receptors. However, the function of Jak in bone metabolism remains to be elucidated. Here, we showed that the inhibition of Jak1 and/or Jak2 in osteoblast-lineage cells led to impaired osteoclastogenesis due to the reduced expression of receptor activator of nuclear factor-κB ligand (RANKL). Murine calvaria-derived osteoblasts induced differentiation of bone marrow cells into osteoclasts in the presence of 1,25-dihydroxyvitamin D3 (1,25D3) and prostaglandin E2 (PGE2) in vitro. However, treatment with the Jak1/2 inhibitor, baricitinib, markedly inhibited osteoclastogenesis in the co-culture. On the other hand, baricitinib did not inhibit RANKL-induced osteoclast differentiation of bone marrow macrophages. These results indicated that baricitinib acted on osteoblasts, but not on bone marrow macrophages. Baricitinib suppressed 1,25D3 and PGE2-induced up-regulation of RANKL in osteoblasts, but not macrophage colony-stimulating factor expression. Moreover, the addition of recombinant RANKL to co-cultures completely rescued baricitinib-induced impairment of osteoclastogenesis. shRNA-mediated knockdown of Jak1 or Jak2 also suppressed RANKL expression in osteoblasts and inhibited osteoclastogenesis. Finally, cytokine array revealed that 1,25D3 and PGE2 stimulated secretion of interleukin-6 (IL-6), IL-11, and leukemia inhibitory factor in the co-culture. Hence, Jak1 and Jak2 represent novel therapeutic targets for osteoporosis as well as inflammatory bone diseases including rheumatoid arthritis. [ABSTRACT FROM AUTHOR]

Prior GWAS have identified loci associated with red blood cell (RBC) traits in populations of European, African, and Asian ancestry. These studies have not included individuals with an Amerindian ancestral background, such as Hispanics/Latinos, nor evaluated the full spectrum of genomic variation beyond single nucleotide variants. Using a custom genotyping array enriched for Amerindian ancestral content and 1000 Genomes imputation, we performed GWAS in 12,502 participants of Hispanic Community Health Study and Study of Latinos (HCHS/SOL) for hematocrit, hemoglobin, RBC count, RBC distribution width (RDW), and RBC indices. Approximately 60% of previously reported RBC trait loci generalized to HCHS/SOL Hispanics/Latinos, including African ancestral alpha- and beta-globin gene variants. In addition to the known 3.8kb alpha-globin copy number variant, we identified an Amerindian ancestral association in an alpha-globin regulatory region on chromosome 16p13.3 for mean corpuscular volume and mean corpuscular hemoglobin. We also discovered and replicated three genome-wide significant variants in previously unreported loci for RDW (SLC12A2 rs17764730, PSMB5 rs941718), and hematocrit (PROX1 rs3754140). Among the proxy variants at the SLC12A2 locus we identified rs3812049, located in a bi-directional promoter between SLC12A2 (which encodes a red cell membrane ion-transport protein) and an upstream anti-sense long-noncoding RNA, LINC01184, as the likely causal variant. We further demonstrate that disruption of the regulatory element harboring rs3812049 affects transcription of SLC12A2 and LINC01184 in human erythroid progenitor cells. Together, these results reinforce the importance of genetic study of diverse ancestral populations, in particular Hispanics/Latinos. [ABSTRACT FROM AUTHOR]

Mast cell (MC) activation contributes considerably to immune responses, such as host protection and allergy. Cell surface immunoreceptors expressed on MCs play an important role in MC activation. Although various immunoreceptors on MCs have been identified, the regulatory mechanism of MC activation is not fully understood. To understand the regulatory mechanisms of MC activation, we used gene expression analyses of human and mouse MCs to identify a novel immunoreceptor expressed on MCs. We found that Tek, which encodes Tie2, was preferentially expressed in the MCs of both humans and mice. However, Tie2 was not detected on the cell surface of the mouse MCs of the peritoneal cavity, ear skin, or colon lamina propria. In contrast, it was expressed on mouse bone marrow–derived MCs and bone marrow MC progenitors (BM-MCps). Stimulation of Tie2 by its ligand angiopoietin-1 induced tyrosine phosphorylation of Tie2 in MEDMC-BRC6, a mouse embryonic stem cell-derived mast cell line, and enhanced MEDMC-BRC6 and mouse BM-MCp adhesion to vascular cell adhesion molecule-1 (VCAM-1) through α4β1 integrin. These results suggest that Tie2 signaling induces α4β1 integrin activation on BM-MCps for adhesion to VCAM-1. [ABSTRACT FROM AUTHOR]

Transfusion of red blood cells (RBCs) is a standard and indispensable therapy in current clinical practice. In vitro production of RBCs offers a potential means to overcome a shortage of transfusable RBCs in some clinical situations and also to provide a source of cells free from possible infection or contamination by microorganisms. Thus, in vitro production of RBCs may become a standard procedure in the future. We previously reported the successful establishment of immortalized mouse erythroid progenitor cell lines that were able to produce mature RBCs very efficiently. Here, we have developed a reliable protocol for establishing immortalized human erythroid progenitor cell lines that are able to produce enucleated RBCs. These immortalized cell lines produce functional hemoglobin and express erythroid-specific markers, and these markers are upregulated following induction of differentiation in vitro. Most importantly, these immortalized cell lines all produce enucleated RBCs after induction of differentiation in vitro, although the efficiency of producing enucleated RBCs remains to be improved further. To the best of our knowledge, this is the first demonstration of the feasibility of using immortalized human erythroid progenitor cell lines as an ex vivo source for production of enucleated RBCs. [ABSTRACT FROM AUTHOR]

Induced pluripotent stem (iPS) cells can be established from somatic cells. However, there is currently no established strategy to generate corneal epithelial cells from iPS cells. In this study, we investigated whether corneal epithelial cells could be differentiated from iPS cells. We tested 2 distinct sources: human adult dermal fibroblast (HDF)-derived iPS cells (253G1) and human adult corneal limbal epithelial cells (HLEC)-derived iPS cells (L1B41). We first established iPS cells from HLEC by introducing the Yamanaka 4 factors. Corneal epithelial cells were successfully induced from the iPS cells by the stromal cell-derived inducing activity (SDIA) differentiation method, as Pax+/K12+ corneal epithelial colonies were observed after prolonged differentiation culture (12 weeks or later) in both the L1B41 and 253G1 iPS cells following retinal pigment epithelial and lens cell induction. Interestingly, the corneal epithelial differentiation efficiency was higher in L1B41 than in 253G1. DNA methylation analysis revealed that a small proportion of differentially methylated regions still existed between L1B41 and 253G1 iPS cells even though no significant difference in methylation status was detected in the specific corneal epithelium-related genes such as K12, K3, and Pax6. The present study is the first to demonstrate a strategy for corneal epithelial cell differentiation from human iPS cells, and further suggests that the epigenomic status is associated with the propensity of iPS cells to differentiate into corneal epithelial cells. [ABSTRACT FROM AUTHOR]

The generation of induced pluripotent stem cells (iPSCs) by introducing reprogramming factors into somatic cells is a promising method for stem cell therapy in regenerative medicine. Therefore, it is desirable to develop a minimally invasive simple method to create iPSCs. In this study, we generated human nasal epithelial cells (HNECs)-derived iPSCs by gene transduction with Sendai virus (SeV) vectors. HNECs can be obtained from subjects in a noninvasive manner, without anesthesia or biopsy. In addition, SeV carries no risk of altering the host genome, which provides an additional level of safety during generation of human iPSCs. The multiplicity of SeV infection ranged from 3 to 4, and the reprogramming efficiency of HNECs was 0.08--0.10%. iPSCs derived from HNECs had global gene expression profiles and epigenetic states consistent with those of human embryonic stem cells. The ease with which HNECs can be obtained, together with their robust reprogramming characteristics, will provide opportunities to investigate disease pathogenesis and molecular mechanisms in vitro, using cells with particular genotypes. [ABSTRACT FROM AUTHOR]

Background: Mesothelioma is a highly malignant tumor that is primarily caused by occupational or environmental exposure to asbestos fibers. Despite worldwide restrictions on asbestos usage, further cases are expected as diagnosis is typically 20- 40 years after exposure. Once diagnosed there is a very poor prognosis with a median survival rate of 9 months. Considering this the development of early pre clinical diagnostic markers may help improve clinical outcomes. Methodology: Microarray expression arrays on mesothelium and other tissues dissected from mice were used to identify candidate mesothelial lineage markers. Candidates were further tested by qRTPCR and in-situ hybridization across a mouse tissue panel. Two candidate biomarkers with the potential for secretion, uroplakin 3B (UPK3B), and leucine rich repeat neuronal 4 (LRRN4) and one commercialized mesothelioma marker, mesothelin (MSLN) were then chosen for validation across a panel of normal human primary cells, 16 established mesothelioma cell lines, 10 lung cancer lines, and a further set of 8 unrelated cancer cell lines. Conclusions: Within the primary cell panel, LRRN4 was only detected in primary mesothelial cells, but MSLN and UPK3B were also detected in other cell types. MSLN was detected in bronchial epithelial cells and alveolar epithelial cells and UPK3B was detected in retinal pigment epithelial cells and urothelial cells. Testing the cell line panel, MSLN was detected in 15 of the 16 mesothelioma cells lines, whereas LRRN4 was only detected in 8 and UPK3B in 6. Interestingly MSLN levels appear to be upregulated in the mesothelioma lines compared to the primary mesothelial cells, while LRRN4 and UPK3B, are either lost or down-regulated. Despite the higher fraction of mesothelioma lines positive for MSLN, it was also detected at high levels in 2 lung cancer lines and 3 other unrelated cancer lines derived from papillotubular adenocarcinoma, signet ring carcinoma and transitional cell carcinoma. [ABSTRACT FROM AUTHOR]

Background. The supply of transfusable red blood cells (RBCs) is not sufficient in many countries. If erythroid cell lines able to produce transfusable RBCs in vitro were established, they would be valuable resources. However, such cell lines have not been established. To evaluate the feasibility of establishing useful erythroid cell lines, we attempted to establish such cell lines from mouse embryonic stem (ES) cells. Methodology/Principal Findings. We developed a robust method to obtain differentiated cell lines following the induction of hematopoietic differentiation of mouse ES cells and established five independent hematopoietic cell lines using the method. Three of these lines exhibited characteristics of erythroid cells. Although their precise characteristics varied, each of these lines could differentiate in vitro into more mature erythroid cells, including enucleated RBCs. Following transplantation of these erythroid cells into mice suffering from acute anemia, the cells proliferated transiently, subsequently differentiated into functional RBCs, and significantly ameliorated the acute anemia. In addition, we did not observe formation of any tumors following transplantation of these cells. Conclusion/Significance. To the best of our knowledge, this is the first report to show the feasibility of establishing erythroid cell lines able to produce mature RBCs. Considering the number of human ES cell lines that have been established so far, the intensive testing of a number of these lines for erythroid potential may allow the establishment of human erythroid cell lines similar to the mouse erythroid cell lines described here. In addition, our results strongly suggest the possibility of establishing useful cell lines committed to specific lineages other than hematopoietic progenitors from human ES cells. [ABSTRACT FROM AUTHOR]

Although nitric oxide (NO) is a known factor that regulates the bone physiology, few and discordant results have been obtained in human studies evaluating the effect of nitrates on bone health. We investigated for the relationship between serum NOx level and incident osteoporotic fracture rate prospectively in a cohort consisting of Japanese women. A total of 871 subjects (67.5 ± 10.8 y/o) were analyzed. During the observation period (8.8 ± 7.2 yrs), incident osteoporotic fractures occurred in 267 participants (209 vertebral fractures, 57 long-bone fractures, and 1 both types). Hazard ratio, by the Cox proportional hazards model, of serum NOx for incident fracture was 0.64 (95% confidence interval 0.53-0.78, p < 0.001) after adjustment for baseline age (1.13, 1.06-1.21, p < 0.001), lumbar bone mineral density (L-BMD; 0.85, 0.78-0.92, p < 0.001), presence of prevalent fracture (3.27, 2.49-4.32, p < 0.001), and treatment of osteoporosis (0.70, 0.53-0.92, p = 0.010). The relationships between serum level of NOx and bone-related parameters were examined by multiple regression analysis; body mass index (p < 0.001) and L-BMD (p = 0.011) were significantly associated with serum NOx level. These results suggest that the low circulating NOx is one of the independent predictors for osteoporotic fracture occurrence in postmenopausal women., Competing Interests: The authors have declared that no competing interests exist., (Copyright: © 2023 Shiraki 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.)