Mesenchymal stem cell (MSC) exosomes promote tissue regeneration and repair, and thus might be used to treat many diseases; however, the influence of microenvironmental conditions on exosomes remains ...unclear. The present study aimed to analyze the effect of osteogenic induction on the functions of human umbilical cord MSC (HucMSC)-derived exosomes. Exosomes from standardized stem cell culture (Exo1) and osteogenic differentiation-exosomes (Exo2) were co-cultured with osteoblasts, separately. Cell counting kit-8 assays, alkaline phosphatase and alizarin red staining were used to observe the exosomes’ effects on osteoblast proliferation and differentiation. The levels of osteogenic differentiation-related proteins were analyzed using western blotting. Estrogen-deficient osteoporosis model mice were established, and treated with the two exosome preparations. Micro-computed tomography and hematoxylin and eosin staining were performed after 6 weeks. MicroRNAs in Exo1 and Exo2 were sequenced and analyzed using bioinformatic analyses. Compared with Exo1 group, Exo2 had a stronger osteogenic differentiation promoting effect, but a weaker proliferation promoting effect. In ovariectomy-induced osteoporosis mice, both Exo1 and Exo2 improved the tibial density and reversed osteoporosis in vivo. High-throughput microRNA sequencing identified 221 differentially expressed microRNAs in HucMSC-derived exosomes upon osteogenic induction as compared with the untreated control group. Importantly, we found that 41 of these microRNAs are potentially critical for MSC-secreted exosomes during osteogenic induction. Mechanistically, exosomal miRNAs derived from osteogenic induced-HucMSCs are involved in bone development and differentiation, such as osteoclast differentiation and the MAPK signaling pathway. The expression of hsa-mir-2110 and hsa-mir-328-3p gradually increased with prolonged osteogenic differentiation and regulated target genes associated with bone differentiation, suggesting that they are probably the most important osteogenesis regulatory microRNAs in exosomes. In conclusion, we examined the contribution of osteogenic induction to the function of exosomes secreted by HucMSCs following osteogenic differentiation in vitro and in vivo, and reveal the underlying molecular mechanisms of exosome action during osteoporosis.
Abnormal accumulation of acyl-CoA cholesterol acyltransferase-1 (ACAT-1) mediated cholesterol ester has been shown to contribute to cancer progression in various cancers including leukemia, glioma, ...breast, pancreatic and prostate cancers. However, the significance of ACAT-1 and cholesterol esters (CE) is relatively understudied in ovarian cancer. In this in vitro study, we assessed the expression and contribution of ACAT-1 in ovarian cancer progression. We observed a significant increase in the expression of ACAT-1 and CE levels in a panel of ovarian cancer cell lines (OC-314, SKOV-3 and IGROV-1) compared to primary ovarian epithelial cells (normal controls). To confirm the tumor promoting capacity of ACAT-1, we inhibited ACAT-1 expression and activity by treating our cell lines with an ACAT inhibitor, avasimibe, or by stable transfection with ACAT-1 specific short hairpin RNA (shRNA). We observed significant suppression of cell proliferation, migration and invasion in ACAT-1 knockdown ovarian cancer cell lines compared to their respective controls (cell lines transfected with scrambled shRNA). ACAT-1 inhibition enhanced apoptosis with a concurrent increase in caspases 3/7 activity and decreased mitochondrial membrane potential. Increased generation of reactive oxygen species (ROS) coupled with increased expression of p53 may be the mechanism(s) underlying pro-apoptotic action of ACAT-1 inhibition. Additionally, ACAT-1 inhibited ovarian cancer cell lines displayed enhanced chemosensitivity to cisplatin treatment. These results suggest ACAT-1 may be a potential new target for the treatment of ovarian cancer.
Osteoporosis is a condition associated with osteolytic bone disease that is primarily characterized by inordinate osteoclast activation. Protein kinase B (Akt) pathways activated by receptor ...activator of nuclear factor kappa-B ligand (RANKL) are essential for osteoclastogenesis. Asiatic acid (AA) is a natural pentacyclic triterpenoid compound extracted from a traditional Chinese herb that exhibits a wide range of biological activities. AA has been found to alleviate the hypertrophic and fibrotic phenotype of chondrocytes via the Akt signaling pathway. In this study, we investigated whether AA alleviated bone loss by inhibiting the Akt signaling pathway during osteoclastogenesis and its effect on osteoblasts. The effect of AA cytotoxicity on mouse bone marrow-derived macrophages/monocytes (BMMs) was evaluated
using a Cell Counting Kit-8 assay. The effects of AA on osteoclast differentiation and function were detected using tartrate-resistant acid phosphatase (TRAP) staining and a pit formation assay. A Western blot and qRT-PCR were conducted to evaluate the expression of osteoclast-specific genes and protein signaling molecules. In addition, alkaline phosphatase and alizarin red staining were performed to assess osteoblast differentiation and mineralization. The bone protective effect of AA was investigated
using ovariectomized mice. we found that AA could dose-dependently inhibit RANKL-induced osteoclastogenesis. Moreover, the pit formation assay revealed that osteoclast function was suppressed by treatment with AA. Moreover, the expression of osteoclast-specific genes was found to be substantially decreased during osteoclastogenesis. Analysis of the molecular mechanisms showed that AA could inhibit NF-kappaB/MAPK/Akt signaling pathway, as well as the downstream factors of NFATc1 in the osteoclast signaling pathway activated by RANKL. However, AA did not significantly promote osteoblast differentiation and mineralization. The
experiments suggested that AA could alleviate ovariectomy-induced bone loss in ovariectomized mice. Our results demonstrate that AA can inhibit osteoclastogenesis and prevent ovariectomy-induced bone loss by inhibiting the NF-kappaB/MAPK/Akt signaling pathway. The discovery of the new molecular mechanism that AA inhibits osteoclastogenesis provides essential evidence to support the use of AA as a potential drug for the treatment of osteoclast-related diseases.
Alveolar Rhabdomyosarcoma is a profoundly malignant soft-tissue sarcoma that predominantly affects children and adolescents. However, the medical field lacks consensus regarding the optimal surgical ...approach to be undertaken in cases where this tumor causes local bone destruction in the upper limb.
A 17-year-old male presented a mass in his left forearm and CT and MRI indicated that the mass had penetrated the ulnar cortex and infiltrating the medulla, resulting in the formation of an eccentric trans-ventricular tumor focus. The sizable tumor affected the volar muscles of the forearm as well as the ulnar bone marrow, exerting pressure on the ulnar artery and vein. It was confirmed by needle biopsy that the mass is alveolar rhabdomyosarcoma. Following two courses of neoadjuvant chemotherapy, the tumor was widely excised en bloc. Autologous fibula with a vascular pedicle was utilized for reconstruction during the procedure. In the postoperative follow-up, no local recurrence of the tumor was observed. Furthermore, the patient retained satisfactory wrist flexion and pronation function in the left forearm.
Alveolar rhabdomyosarcoma is an uncommon and highly aggressive form of soft tissue sarcoma. Scientific management necessitates a multidisciplinary approach, combining chemotherapy with surgery. In cases where the tumor invaded into compartment of the bone, careful consideration should be given to the boundaries of tumor resection, the extent of osteotomy, and the approach to musculoskeletal reconstruction when designing the surgical plan. Through reporting our own case and thoroughly reviewing previous clinical experiences, we aim to provide valuable insights for the treatment of this particular disease.
Abnormal accumulation of acyl-CoA cholesterol acyltransferase-1 (ACAT1) and ACAT1-mediated cholesterol esterified with fatty acids (CE) contribute to cancer progression in various cancers. Our ...findings of increased CE and ACAT1 levels in epithelial ovarian cancer (EOC) cell lines prompted us to investigate whether such an increase occurs in primary clinical samples obtained from human subjects diagnosed with EOC. We evaluated the diagnostic/prognostic potential of ACAT1 and CE in EOC by: 1) assessing ACAT1 and CE levels in plasma, peritoneal fluid, and ovarian/tumor tissues; 2) assessing diagnostic performance by Receiver Operating Characteristic (ROC) analysis; and 3) comparing expression of ACAT1 and CE with that of tumor proliferation marker, Ki67.
ACAT1 protein levels in plasma, peritoneal fluid and tissue were measured via enzyme-linked immunosorbent assay. Tissue expression of ACAT1 and Ki67 proteins were confirmed by immunohistochemistry and mRNA transcript levels were evaluated using quantitative real-time polymerase chain reaction (qRT-PCR). CE levels were assessed in plasma, peritoneal fluid (colorimetric assay) and in tissue (thin layer chromatography).
Preoperative levels of ACAT1 and CE on the day of surgery were significantly higher in tissue and peritoneal fluid from EOC patients vs. the non-malignant group, which included subjects with benign tumors and normal ovaries; however, no significant differences were observed in plasma. In tissue and peritoneal fluid, positive correlations were observed between CE and ACAT1 levels, as well as between ACAT1/CE and Ki67.
ACAT1 and CE accumulation may be linked to the aggressive potential of EOC; therefore, these mediators may be useful biomarkers for EOC prognosis and target-specific treatments.
The efficacy of immune checkpoint inhibitors (ICIs), including toripalimab and pembrolizumab, has not been confirmed in the treatment of cancer of unknown primary (CUP), which has a very poor ...prognosis. Combined with anti-angiogenic therapies, ICIs are hypothesized to be effective in prolonging overall survival. The study aims to give evidence on the treatment effects of sunitinib combined with ICIs, find pathological biomarkers associated with changes in volumetric
F FDG PET/CT parameters, and investigate inner associations among these markers associated with response on PET/CT.
The study recruited patients receiving combined treatment (ICIs + sunitinib), compared the effects of combined treatment with those of separate treatment and age-matched negative controls, and analyzed propensity score-matched (PSM) pairs. Markers associated with survival were identified, and their inner associations were tested using structural equation modeling.
A total of 292 patients were enrolled in the final analysis, with 53 patients receiving combined treatment. Survival analysis demonstrated significantly prolonged survival in either combined or separate treatment, with the combined arm showing better response when PSM-paired using pre-treatment whole-body PET/CT parameters. The angiogenic markers KDR and VEGF mediate the PD-1 blockade impact on volumetric value changes in positive and negative manners.
The anti-angiogenic agent sunitinib may potentiate PD-1 blockade by diminishing angiogenesis or its downstream effects. The combined separate treatment increased the survival of CUP patients, and the responses could be evaluated using volumetric PET/CT parameters.
The Pseudomonas aeruginosa strain, PAO1, has three putative γ-glutamyltranspeptidase (GGT) genes: ggtI, ggtII, and ggtIII. In this study, the expression of each of these genes in P. aeruginosa PAO1 ...was analyzed, and the properties of the corresponding GGT proteins were investigated. This is the first report on biochemical characterization of GGT paralogs from Pseudomonas species. The crude extracts prepared from P. aeruginosa PAO1 exhibited hydrolysis and transpeptidation activities of 17.3 and 65.0 mU/mg, respectively, and the transcription of each gene to mRNA was confirmed by RT-PCR. All genes were cloned, and the expression plasmids constructed were introduced into an Escherichia coli expression system. Enzyme activity of the expressed protein of ggtI (PaGGTI) was not detected in the system, while the enzyme activities of the expressed proteins derived from ggtII and ggtIII (PaGGTII and PaGGTIII, respectively) were detected. However, the enzyme activity of PaGGTII was very low and easily decreased. PaGGTII with C-terminal his-tag (PaGGTII25aa) showed increased activity and stability, and the purified enzyme consisted of a large subunit of 40 kDa and a small subunit of 28 kDa. PaGGTIII consisted of a large subunit of 37 kDa and a small subunit of 24 kDa. The maximum hydrolysis and transpeptidation activities of PaGGTII25aa were obtained at 40ºC–50ºC, and the maximum hydrolysis and transpeptidation activities of PaGGTIII were obtained at 50ºC–60ºC. These enzymes retained approximately 80% of their hydrolysis and transpeptidation activities after incubation at 50ºC for 10 min, reflecting good stability. Both PaGGTII25aa and PaGGTIII showed higher activities of hydrolysis and transpeptidation in the alkali range than in the acidic range. However, they were highly stable at a wide pH range (5–10.5).
Although there is extended research on the response to severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) vaccines in adult cancer patients (ACP), the immunogenicity to the variants of ...concern (VOCs) in childhood cancer patients (CCP) and safety profiles are now little known.
A prospective, multi-center cohort study was performed by recruiting children with a solid cancer diagnosis and childhood healthy control (CHC) to receive standard two-dose SARS-CoV-2 vaccines. An independent ACP group was included to match CCP in treatment history. Humoral response to six variants was performed and adverse events were followed up 3 months after vaccination. Responses to variants were compared with ACP and CHC by means of propensity score-matched (PSM) analysis.
The analysis included 111 CCP (27.2%, median age of 8, quartile 5.5-15 years), 134 CHC (32.8%), and 163 ACP (40.0%), for a total 408 patients. Pathology included carcinoma, neural tumors, sarcoma, and germ cell tumors. Median chemotherapy time was 7 (quartile, 5-11) months. In PSM sample pairs, the humoral response of CCP against variants was significantly decreased, and serology titers (281.8 ± 315.5 U/ml) were reduced, as compared to ACP (
< 0.01 for the rate of neutralization rate against each variant) and CHC (
< 0.01 for the rate of neutralization against each variant) groups. Chemotherapy time and age (Pearson
≥ 0.8 for all variants) were associated with the humoral response against VOCs of the CHC group. In the CCP group, less than grade II adverse events were observed, including 32 patients with local reactions, and 29 patients had systemic adverse events, including fever (
= 9), rash (
= 20), headache (
= 3), fatigue (
= 11), and myalgia (
= 15). All reactions were well-managed medically.
The humoral response against VOCs after the CoronaVac vaccination in CCP was moderately impaired although the vaccine was safe. Age and chemotherapy time seem to be the primary reason for poor response and low serology levels.
Severe acute respiratory syndrome-coronavirus 2 (COVID-19) vaccines may incur changes in thyroid functions followed by mood changes, and patients with Hashimoto thyroiditis (HT) were suggested to ...bear a higher risk.
We primarily aim to find whether COVID-19 vaccination could induce potential subsequent thyroid function and mood changes. The secondary aim was to find inflammatory biomarkers associated with risk.
The retrospective, multi-center study recruited patients with HT receiving COVID-19-inactivated vaccines. C-reactive proteins (CRPs), thyroid-stimulating hormones (TSHs), and mood changes were studied before and after vaccination during a follow-up of a 6-month period. Independent association was investigated between incidence of mood state, thyroid functions, and inflammatory markers. Propensity score-matched comparisons between the vaccine and control groups were carried out to investigate the difference.
Final analysis included 2,765 patients with HT in the vaccine group and 1,288 patients in the control group. In the matched analysis, TSH increase and mood change incidence were both significantly higher in the vaccine group (11.9% versus 6.1% for TSH increase and 12.7% versus 8.4% for mood change incidence). An increase in CRP was associated with mood change (p< 0.01 by the Kaplan-Meier method) and severity (r = 0.75) after vaccination. Baseline CRP, TSH, and antibodies of thyroid peroxidase (anti-TPO) were found to predict incidence of mood changes.
COVID-19 vaccination seemed to induce increased levels and incidence of TSH surge followed by mood changes in patients with HT. Higher levels of pre-vaccine serum TSH, CRP, and anti-TPO values were associated with higher incidence in the early post-vaccine phase.
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