Electrospun Biomimetic Periosteum Capable of Controlled Release of Multiple Agents for Programmed Promoting Bone Regeneration.

The effective repair of large bone defects remains a major challenge due to its limited self-healing capacity. Inspired by the structure and function of the natural periosteum, an electrospun biomimetic periosteum is constructed to programmatically promote bone regeneration using natural bone healing mechanisms. The biomimetic periosteum is composed of a bilayer with an asymmetric structure in which an aligned electrospun poly(ε-caprolactone)/gelatin/deferoxamine (PCL/GEL/DFO) layer mimics the outer fibrous layer of the periosteum, while a random coaxial electrospun PCL/GEL/aspirin (ASP) shell and PCL/silicon nanoparticles (SiNPs) core layer mimics the inner cambial layer. The bilayer controls the release of ASP, DFO, and SiNPs to precisely regulate the inflammatory, angiogenic, and osteogenic phases of bone repair. The random coaxial inner layer can effectively antioxidize, promoting cell recruitment, proliferation, differentiation, and mineralization, while the aligned outer layer can promote angiogenesis and prevent fibroblast infiltration. In particular, different stages of bone repair are modulated in a rat skull defect model to achieve faster and better bone regeneration. The proposed biomimetic periosteum is expected to be a promising candidate for bone defect healing.

Application of Three-Dimension Printing Nano-Carbonated-Hydroxylapatite to the Repair of Defects in Rabbit Bone.

Introduction: Hydroxylapatite (HAp) is a biodegradable bone graft material with high biocompatibility. However, the clinical application of HAp has been limited due to the poor absorption rate in vivo. Methods: In this study, carbonated hydroxylapatite (CHAp) with a chemical composition similar to natural bone was synthesized. HAp and CHAp scaffolds were fabricated by 3D printing. Each material was designed by two types of scaffold model with a maximum width of 8 mm and a thickness of 2 mm, ie, structure I (round shape) and structure II (grid shape). Then, the HAp scaffolds were loaded with lutein. These scaffolds were implanted into the 8 mm bone defect on the top of the rabbit skull within 3 hours in the morning. The curative effects of the scaffolds were assessed two months after implantation. Results: The 3D printed scaffolds did not cause severe inflammation or rejection after implantation. It showed that the porous structures allow bone cells to enter into the scaffolds. Furthermore, CHAp scaffolds were more biocompatible than HAp scaffolds, and showed a higher level of degradation and new bone formation after implantation. Structure II scaffolds with a smaller mineral content degraded faster than structure I, while structure I had better osteoconductive properties than structure II. Besides, the addition of lutein significantly enhanced the rate of new bone formation. Discussion: The uniqueness of this study lies in the synthesis of 3D printed CHAp scaffolds and the implantation of CHAp in rabbit bone defects. The incorporation of suitable carbonate and lutein into HAp can enhance the osteoinductivity of the graft, and CHAp has a faster degradation rate in vivo, all of which provide a new reference for the research and application of apatite-based composites.

Coated sodium butyrate and vitamin D3 supplementation improve gut health through influencing intestinal immunity, barrier, and microflora in early-stage broilers.

BACKGROUND: Intestinal development and function are critical to maintaining sustained broiler growth. The present study aimed to evaluate the effects of coated sodium butyrate (CSB) and vitamin D3 (VD3) on the intestinal immunity, barrier, oxidative stress and microflora in early-stage broilers. In total, 192 one-day-old broilers were assigned to a 2 × 2 factorial design including two dietary supplements at two different levels, in which the main effects were VD3 (3000 or 5000 IU kg-1) and CSB (0 or 1 g kg-1). RESULTS: The results showed that CSB supplementation increased ileal goblet cells (GCs) numbers, villus height and decreased crypt depth in broilers. CSB increased ileal proliferating cell nuclear antigen expression and high-level VD3 decreased cluster of differentiation 3 expression. CSB reduced serum d-lactate, endotoxin (ET), adrenocorticotropic hormone, corticosterone and malondialdehyde (MDA) concentrations and increased total antioxidant capacity (T-AOC) level. Meanwhile, high-level VD3 decreased serum ET concentration. Furthermore, CSB increased ileal T-AOC, lysozyme (LYZ) and transforming growth factor (TGF)-ß and decreased MDA, whereas high-level VD3 decreased ileal MDA and increased secretory immunoglobulin A. CSB up-regulated ileal claudin1, superoxide dismutase 1, TGF-ß and LYZ mRNA expression and down-regulated interleukin-1ß mRNA expression. CSB combined with high-level VD3 increased ileal Faecalibaculum abundance. Spearman correlation analysis showed that Faecalibaculum was related to the immune and barrier function. CONCLUSION: Dietary supplementation with CSB and high-level VD3 improved early gut health in broilers by promoting intestinal development, enhancing antioxidant capacity, strengthening barrier function and enhancing the favorable composition of the gut bacterial flora. © 2024 Society of Chemical Industry.

Chitosan nonwoven fabric composited calcium alginate and adenosine diphosphate as a hemostatic bandage for acute bleeding wounds.

Acute bleeding following accidental injury is a leading cause of mortality. However, conventional hemostatic bandages impede wound healing by inducing excessive blood loss, dehydration, and adherence to granulation tissue. Strategies such as incorporating active hemostatic agents and implementing chemical modifications can augment the properties of these bandages. Nevertheless, the presence of remote thrombosis and initiators may pose risks to human health. Here, a hemostatic bandage was developed by physically combined chitosan nonwoven fabric, calcium alginate sponge, and adenosine diphosphate. The presented hemostatic bandage not only exhibits active and passive mechanisms for promoting clotting but also demonstrates excellent mechanical properties, breathability, ease of removal without causing damage to the wound bed or surrounding tissues, as well as maintaining an optimal moist environment conducive to wound healing. In vitro evaluation results indicated that the hemostatic bandage possesses favorable cytocompatibility with low levels of hemolysis. Furthermore, it effectively aggregates various blood cells while activating platelets synergistically to promote both extrinsic and intrinsic coagulation pathways. In an in vivo rat model study involving liver laceration and femoral artery injury scenarios, our developed hemostatic bandage demonstrated rapid clot formation capabilities along with reduced blood loss compared to commercially available fabrics.

Biomechanical and tissue reaction: the effects of varying sutures size on canine abdominal wall stitching.

Objective: Larger diameter sutures can provide sufficient tensile strength to surgical incisions but may exacerbate the inflammatory response caused by the amount of implanted foreign material. This experiment aims to investigate the differences in biomechanical stability and tissue reactivity after suturing canine midline abdominal incisions with different suture sizes. Method: Assessing the biomechanical differences between USP 2-0, 3-0, and 4-0 PGA sutures using uniaxial tensile testing on ex vivo canine midline skin and fascial muscle tissues using either a simple continuous or simple interrupted technique. mRNA and protein expression levels of inflammatory factors were measured through RT-PCR and ELISA. Tissue reactivity was evaluated using a semi-quantitative scoring system. Result: For strains below 30% in skin and below 50% in muscle, there were no significant differences among groups. The results of skin biomechanical testing showed that the USP 4-0 PGA suture group demonstrated significantly lower maximum tensile strength compared to the USP 2-0 PGA or USP 3-0 PGA suture groups. However, it remained capable of providing at least 56.3 N (1.03 MPa) tensile strength for canine skin incisions, matching the tensile strength requirements of general canine abdominal wall surgical incisions. In addition, there were no statistically significant differences observed in the maximum tensile strength among different size of sutures according to the data of biomechanical testing in muscle. Larger diameter sutures led to increased levels of inflammatory factors (IL-1ß, IL-6, TNF-ɑ) and tissue reactivity. Simple interrupted sutures caused higher levels of inflammatory factors in muscular tissue compared to simple continuous sutures. Conclusion: USP 4-0 PGA sutures provide sufficient biomechanical stability for suturing canine abdominal skin and linea alba. Suture size significantly influences tissue reactivity after suturing, with smaller gauge sutures reducing early tissue inflammatory response. Thus, USP 4-0 PGA suture has more advantages to suturing canine abdominal surgical incisions.

Alfaxalone Alleviates Neurotoxicity and Cognitive Impairment Induced by Isoflurane Anesthesia in Offspring Rats.

BACKGROUND: The anesthetic isoflurane can cause neurotoxicity in fetuses and offspring of rats, affecting their neurodevelopment. However, the underlying mechanisms and therapeutic targets of isoflurane-induced neurotoxicity remain to be identified. Alfaxalone (ALF) is a steroid anesthetic. Steroids have been reported to have neuroprotective effects. This study aimed to investigate whether ALF could alleviate the isoflurane-induced neurotoxicity in fetuses and offspring of rats. METHODS: On gestation day 15 (G15), the pregnant SD rats were randomly assigned to 4 groups: control 1 (CTL1) + control 2 (CTL2), isoflurane (ISO) + CTL2, CTL1 + ALF, and ISO + ALF. To analyze the changes in the expression levels of inflammatory cytokines, apoptotic factors, and synaptophysin, the brain tissues from the G15 fetuses and offspring at postnatal day 7 (P7), postnatal day 14 (P14), and postnatal day 31 (P31) were collected. The newborn neurons in the rats' offspring at P7, P14, and P31 were counted using immunofluorescence techniques. The Morris water maze (MWM) test was performed to assess the learning and memory abilities of P31 offspring rats. RESULTS: ALF significantly alleviated the isoflurane-induced increase in the expression levels of inflammatory cytokines and apoptotic factors, such as interleukin (IL)-6 (ISO + CTL2 versus ISO + ALF: 5.133 ± 0.739 versus 1.093 ± 0.213, P < .001) and Caspase-3 (6.457 ± 0.6 versus 1.062 ± 0.1, P < .001) in the G15 fetuses. In P31 offspring rats, the expression levels of synaptophysin (0.719 ± 0.04 versus 1.068 ± 0.072, P < .001) and the number of newborn neurons in the dentate gyrus of the hippocampus were significantly lower in the ISO + CTL2 group as compared to those in the ISO + ALF group (118 ± 6 versus 140 ± 7, P < .001). These changes also occurred in the rat offspring at P7 and P14. In the MWM test, the escape latency of CTL1 + ALF group rats was significantly lower than that of ISO + ALF group rats (41 ± 6 versus 31 ± 7, P < .001) at P31. CONCLUSIONS: Based on these findings, this study suggested that isoflurane exposure during pregnancy in rats could cause neuroinflammation and death of embryos as well as impairment of cognitive function in the offspring rats. ALF can be used to counteract the negative effects of isoflurane.

Affecting factors for abdominal incisional tension in surgery of dogs and cats.

OBJECTIVE: Proper assessment of intraoperative abdominal incisional tension helps to select the appropriate sutures and suture method. Wound tension is usually thought to be associated with wound size, but few relevant articles have been reported. The aim of this study was to investigate the core factors influencing abdominal incisional tension and construct regression equations to judge incisional tension in clinical surgery. METHODS: Medical records were collected from clinical surgical cases at the Teaching Animal Hospital of Nanjing Agricultural University from March 2022 to June 2022. The data collected mainly included the body weight, and the incisional length, margin, and tension. The core factors affecting abdominal wall incisional tension were screened by correlation analysis, random forest analysis, and multiple linear regression analysis. RESULTS: Although correlation analysis showed that multiple same and deep layer abdominal incision parameters and body weight were significantly correlated with abdominal incisional tension. However, the same layer of abdominal incisional margin had the largest correlation coefficient. In random forest models, the abdominal incisional margin had the main contribution to the prediction of the same layer's abdominal incisional tension. In the multiple linear regression model, all incisional tension could be predicted by the same layer of abdominal incisional margin as the only independent variable, except for canine muscle and subcutaneous. The canine muscle and subcutaneous incisional tension were binary regressions with the same layer's abdominal incision margin and body weight. CONCLUSION: The same layer's abdominal incisional margin is the core factor positively related to the abdominal incisional tension intraoperatively.

Expression and Regulation of the GABAA Receptor/STEP61 Signaling Pathway in Cerebral Cortical Neurons Treated with Emulsified Isoflurane In Vitro.

Emulsified isoflurane (EISO) is an intravenous anesthetic. However, researchers have not clearly determined how emulsified isoflurane affects the central nervous system during the process of anesthesia. The aim of this study was to explore changes in the gamma-aminobutyric acid type A receptor subunit (GABAA), 61 kD isoform of striatal-enriched protein phosphatase (STEP61) signaling pathway, and epigenetic regulation in cortical neurons after treatment with emulsified isoflurane. After immunological identification, the isolated neurons were randomly divided into three groups: the blank group (Con), intralipid treatment group (FE), and emulsified isoflurane treatment group (EISO). Neuron viability was assayed using cell counting kit-8 (CCK-8). The expression levels of target nucleic acids, proteins, and corresponding ligands were detected. Using real-time polymerase chain reaction (PCR) to assess the promoter methylation of ion channel proteins in the cerebral cortex of rats anesthetized with EISO, we observed changes in promoter methylation of the genes encoding gamma-aminobutyric acid type A receptor α1 subunit (GABAAα1), N-methyl-d-aspartate receptor subunit 1 (NMDAR1), and mu opioid receptor 1 (OPRM1), accompanied by changes in the levels of their messenger ribonucleic acids (mRNAs) and proteins. The levels of ligands for these receptors were also altered. EISO altered the methylation rate of the promoter region of channel protein-coding genes involved in the GABAA/STEP61 signaling pathway in cerebral cortical neurons to regulate gene expression. The ligands for the receptors were also changed.

Camptothecin induced DDX5 degradation increased the camptothecin resistance of osteosarcoma.

Osteosarcoma (OS) is the most common primary malignant bone tumor in children and adolescents. Unfortunately, chemo-resistance is a huge obstacle in the treatment of OS. However, the underlying molecular mechanisms of OS chemo-resistance still remain unknown. Here we reported that the resistance to camptothecin (cpt) therapy was driven by degradation of DDX5. DDX5 knockdown decreased cell death and DNA damage and recovered cell proliferation in cpt treated 143B cells. Furthermore, we found that DDX5 bound to NONO, a kind of DNA repairing protein, and regulated NONO functions. Our data verified that cpt-induced degradation of DDX5 following by breaking down the protein bound of NONO, which participated in the resistance of cpt. In the summary, according to our results, DDX5 might be a potential therapeutic target for improving clinical outcomes of cpt in OS.

GABAA Receptor/STEP61 Signaling Pathway May Be Involved in Emulsified Isoflurane Anesthesia in Rats.

(1) Background: Emulsified isoflurane (EISO) is a type of intravenous anesthetic. How emulsified isoflurane works in the brain is still unclear. The aim of this study was to explore whether epigenetic mechanisms affect anesthesia and to evaluate the anesthetic effects of emulsified isoflurane in rats. (2) Methods: Rats were randomly divided into four groups (n = 8/group): The tail vein was injected with normal saline 0.1 mL·kg-1·min-1for the control (Con) group, with intralipid for the fat emulsion (FE) group, with EISO at 60 mg·kg-1·min-1 for the high-concentration (HD) group, and 45 mg·kg-1·min-1 for the low-concentration (LD) group. The consciousness state, motor function of limbs, and response to nociceptive stimulus were observed after drug administration. (3) Results: Using real-time polymerase chain reaction (PCR) to assess the promoter methylation of ion channel proteins in the cerebral cortex of rats anesthetized by EISO, we demonstrated that the change in the promoters' methylation of the coding genes for gamma-aminobutyric acid A receptor α1 subunit (GABAAα1), N-methyl-D-aspartate receptor subunit 1 (NMDAR1), and mu opioid receptor 1 (OPRM1) was accompanied by the change in messenger ribonucleic acid (mRNA) and protein expression by these genes. (4) Conclusion: These data suggest that the epigenetic factors' modulation might offer a novel approach to explore the anesthetic mechanism of EISO.

Detection of a 5-circRNA signature to improve prognostic prediction in gastric cancer.

Gastric cancer (GC) is a commonly diagnosed malignancy with a high mortality rate worldwide. Despite advances in therapeutic approaches, the 5-year survival rate of patients with GC remains poor. A type of non-coding RNA, circular RNA (circRNA), has been discovered. Some circRNAs are dysregulated in cancer and may have important functions. Thus, circRNAs could be candidate biomarkers for disease diagnosis and prognosis. Our study used a reannotation framework to derive expression values of circRNAs from microarray data sets. We used an integrated pipeline including differential expression analysis and Cox regression analyses to detect GC-associated circRNA signatures. We validated results with 54 paired gastric tumor and adjacent normal tissues. Five circRNA signatures were highly associated with patient overall survival and disease-free survival. Real-time PCR experiments on hsa_circ_0103398 and hsa_circ_0127859 in tumors and normal tissues showed that these circRNAs were significantly upregulated in GC. High expression of hsa_circ_0103398 and hsa_circ_0127859 was closely associated with unfavorable survival time. Our findings indicated that a 5-circRNA signature might serve as a candidate prognostic biomarker of GC.

Differences in pathogenicity and virulence-associated gene expression among Pasteurella multocida strains with high and low virulence in a lung tissue model.

Pasteurella multocida capsular type A can cause a pulmonary infection, leading to serious pecuniary losses in cattle. The heterogeneity of infection outcome of P. multocida strains showing different virulence may be related to divergent expression of virulence genes. In this study, we compared the transcriptional response of virulence-associated genes in high (PMPAN001) and low (PMPAN007) virulence P. multocida capsular type A strains in lung tissues and in vitro. These clinical isolates differ in their organ bacterial loads, mRNA abundance of the same virulence genes between lung and culture medium, and extent of lung damage. Among the eight virulence-associated genes (fimA, tbpA, exbD, fur, oma87, pmHAS, nanH, and tonB), seven genes showed higher expression in lung compared with in vitro at 16 h (P ≤ 0.05) in PMPAN001, but not in PMPAN007. FimA, exbD, fur, oma87, pmHAS, and tonB gene transcripts showed significantly higher expression in PMPAN001 than in PMPAN007 in the lung tissues at 16 h post-infection (P ≤ 0.05). Specially, the virulence gene, nanH, in both strains was associated with poor expression in vitro and lung tissue (mean relative mRNA abundance values < 0.6). Strain PMPAN001 had a higher proliferation rate in vivo than strain PMPAN007. The bacterial loads of PMPAN001 in the organs increased from 12 h post-infection, with maximum bacteria count ranging from 1 million to 20 million/mg. In addition, lungs treated with PMPAN001 produced serious and extensive lesions marked with inflammation at 20 h. Overall, our results reveal that the highly expressed virulence-associated genes, fimA, exbD, fur, oma87, pmHAS, and tonB can be used as markers for assessing the virulence of P. multocida capsular type A strains.

Targeting the FOXM1-regulated long noncoding RNA TUG1 in osteosarcoma.

Long noncoding RNAs (lncRNAs) play an important role in the proliferation and metastasis of osteosarcoma. Identification of the pathogenesis of osteosarcoma and development of new therapeutic strategies against osteosarcoma are urgently needed. In this study, we evaluated the expression of TUG1 (Taurine Upregulated Gene 1) in osteosarcoma tissues and selected it as our target for further analyses. In vitro, we found that TUG1 was upregulated by FOXM1 (Forkhead Box M1) in osteosarcoma cells. TUG1 accelerated osteosarcoma proliferation, migration, and invasion by competitively sponging miR-219a-5p, leading to upregulation of Phosphatidylinositol-4, 5-Bisphosphate 3-Kinase Catalytic Subunit Alpha and activation of the protein kinase B (AKT) signaling pathway. In addition, the AKT pathway activation promoted TUG1 expression by upregulating the expression of FOXM1, forming a positive feedback loop in osteosarcoma. Furthermore, we designed and synthesized therapeutic locked nucleic acids targeting TUG1. The proliferation of osteosarcoma was significantly repressed. Hence, TUG1 may be a potential biomarker and therapeutic target for osteosarcoma.

Letrozole induced low estrogen levels affected the expressions of duodenal and renal calcium-processing gene in laying hens.

Estrogen regulates the calcium homeostasis in hens, but the mechanisms involved are still unclear fully. In this study, we investigated whether letrozole (LZ) induced low estrogen levels affected the calcium absorption and transport in layers. In the duodenum, we observed a significant decrease of mRNA expressions of Calbindin-28k (CaBP-28k) and plasma membrane Ca2+-ATPase (PMCA 1b) while CaBP-28k protein expression was declined in birds with LZ treatment, and the mRNA levels of duodenal transient receptor potential vanilloid 6 (TRPV6) and Na+/Ca2+ exchanger 1 (NCX1) were not affected. Interestingly, we observed the different changes in the kidney. The renal mRNA expressions of TRPV6 and NCX1 were unregulated while the PMCA1b was down-regulated in low estrogen layers, however, the CaBP-28k gene and protein expressions were no changed in the kidney. Furthermore, it showed that the duodenal estradiol receptor 2 (ESR2) transcripts rather than parathyroid hormone 1 receptor (PTH1R) and calcitonin receptor (CALCR) played key roles to down-regulate calcium transport in LZ-treated birds. In conclusion, CaBP-28k, PMCA 1b and ESR2 genes in the duodenum may be primary targets for estrogen regulation in order to control calcium homeostasis in hens.

Change of precipitation characteristics in the water-wind erosion crisscross region on the Loess Plateau, China, from 1958 to 2015.

Precipitation plays an important and crucial role in processes in the water-wind erosion crisscross region of the Loess Plateau than in other parts of the region. We analyzed precipitation data and standardized precipitation index (SPI) at 14 representative synoptic stations from 1958 to 2015 used trend-free prewhitening, linear trend estimation, Spearman's rho test, the Mann-Kendall trend test, the Mann-Kendall abrupt change test and rescaled range analysis. The following conclusions were drawn. First, the analysis of monthly precipitation at all stations suggested that precipitation during the rainy season (July, August, September), especially rain in July and August, exhibited a general decreasing trend, while both increasing and decreasing trends were observed in other months. Moreover, the annual precipitation of all stations continued to exhibit decreasing trends except Wuzhai. Erosive rainfall frequency in the rainy season and the annual scale was weakly reduced but erosive force of single rainfall has been enhanced. Second, the SPI exhibited different increasing degrees in winter, while decreasing trends were observed in other seasons. Additionally, the annual-scale SPI at most stations exhibited a stable and sustained downward trend. Therefore, this region is currently associated with a drought trend, and the drought degree will likely continue to increase.

Climatic and drought characteristics in the loess hilly-gully region of China from 1957 to 2014.

The loess hilly-gully region is a focus region of the "Grain for Green" program in China. Drought is the main problem in the study region. Precipitation and temperature are two indicators that directly characterize climatic drought. A thorough analysis of the precipitation, temperature and drought characteristics of the loess hilly-gully region can clarify the current water and heat conditions in the region to improve regional water resource management and provide a reliable reference for effectively improving water use efficiency. In this study, we fully analyzed the precipitation and temperature characteristics at 11 representative synoptic stations in the loess hilly-gully region over the period from 1957 to 2014 using a combination of trend-free pre-whitening, linear trend estimation, Spearman's rho test, the Mann-Kendall (M-K) trend and abrupt change tests and wavelet analysis. The standardized precipitation evapotranspiration index was calculated and analyzed on different time scales. The following conclusions were drawn: (1) There were significant spatial differences and inter-annual variations in precipitation at the 11 synoptic stations in the study area between 1957 and 2014; the precipitation consistently decreased with fluctuations, and the extent of the decrease varied from a maximum of 17.74 mm/decade to a minimum of 2.92 mm/decade. Except for the downward trends of the autumn and winter mean temperatures at Hequ, the seasonal and annual mean temperatures at the stations showed upward trends, including highly significant upward trends. (2) Alternating drought and wetness occurred in the study area; the wet period mainly appeared in the 1960s, and the main dry period lasted from the late 20th century to 2012. There were fewer dry and wet years than normal years; however, the study area still showed a drying trend, and the severity of the drought was increasing. (3) The annual precipitation and annual mean temperature showed marked cyclical fluctuations at each synoptic station, and the first primary cycle was approximately 28 years. The seasonal precipitation and seasonal temperature showed different cycle lengths; the seasonal cycles of precipitation for spring, summer, autumn and winter were 10, 28, 10 and 26 years long, respectively, and the cycles of the temperature fluctuations for all four seasons were approximately 28 years long.

Paeonol protects against TNF-α-induced proliferation and cytokine release of rheumatoid arthritis fibroblast-like synoviocytes by upregulating FOXO3 through inhibition of miR-155 expression.

BACKGROUND: Fibroblast-like synoviocytes (FLS) play an essential role in the pathogenesis of chronic inflammatory diseases, such as rheumatoid arthritis. Paeonol (Pae) is a phenolic compound found in many traditional Chinese medicine remedies. However, the effects of Pae on TNF-α-stimulated FLS and the underlying molecular mechanism are unknown. In this study, we aimed to investigate the anti-proliferative and anti-inflammatory effect of Pae against activated FLS. MATERIALS AND METHODS: Rheumatoid arthritis FLS (RA-FLS) were pre-treated with different doses (25, 50, and 100 µM) of Pae or miR-155 inhibitor for 30 min or transfected with miR-155 mimic, and then treated with 50 ng/mL of tumor necrosis factor alpha (TNF-α) for 1 h. Cells that were untreated served as control. At 24 h after drug pretreatment, the proliferation of FLS was detected using the MTT assay. The concentrations of interleukin IL-6 and IL-1ß in cell culture supernatant were examined by enzyme-linked immunosorbent assay (ELISA), and mRNA levels of Foxo3 and miR-155 expression in FLS were quantified by reverse transcription-polymerase chain reaction (RT-PCR). Protein expressions of forkhead box O3 (FOXO3), cyclin D1, and c-Myc were detected by Western Blot. RESULTS: TNF-α induced the proliferation of FLS, whereas Pae inhibited this proliferation in a dose-dependent manner. Pae attenuated TNF-α-induced production of IL-6 and IL-1ß, and inhibited the expression of miR-155 in a dose-dependent manner. In addition, miR-155 inhibitor decreased TNF-α-induced proliferation of FLS, and attenuated TNF-α-induced production of IL-6 and IL-1ß. In addition, pretreatment with different doses of Pae or miR-155 inhibitor markedly attenuated TNF-α-induced decrease in protein expression of FOXO3 in FLS. Mechanistic studies revealed FOXO3 as miR-155-5p direct target and inhibition of FOXO3 led to the abolishment of Pae protective effects. CONCLUSIONS: Paeonol protected against TNF-α-induced proliferation and cytokine release of FLS by decreasing the expression of miR-155 and upregulating its target FOXO3.

A polysaccharide from Trametes robiniophila inhibits human osteosarcoma xenograft tumor growth in vivo.

In the present study, we isolated and purified one polysaccharide (TRP) from Trametes robiniophila, which had a backbone of 1,3,6- and 1,4-linked glucpyranosyl moieties, with 1-linked arabinofuranosyl and galactopyranosyl terminal at the O-3 position of 1,3,6-linked glucpyranosyl residues. TRP was further evaluated for its antitumor activity against xenografted U-2 OS osteosarcoma in BALB/c nude mice together with the possible mechanism of action. We found that oral administration of TRP significantly suppressed U-2 OS tumor growth in mice via the induction of apoptosis, as evidenced by the increased number of TUNEL-positive cells in tumor tissues. Moreover, TRP administration increased the levels of the proapoptotic Bax protein and decreased the level of the antiapoptotic Bcl-2 protein, thus resulting in a rise of Bax/Bcl-2 ratio. Furthermore, the protein expression of caspase-9, caspase-3 and cleaved PARP became evident in tumor tissues from mice following TRP treatment, but caspase-8 keep unchanged. Besides, overexpression of metadherin (MTDH) was attenuated in tumor tissues of TRP-fed mice. Taken together, these findings suggest that the TRP-induced apoptosis of tumor tissues is through a mitochondria-mediated intrinsic apoptotic pathway.

A polysaccharide from Trametes robiniophila Murrill induces apoptosis through intrinsic mitochondrial pathway in human osteosarcoma (U-2 OS) cells.

In this study, we isolated and purified one homogeneous polysaccharide (TRP) from the fruiting bodies of Trametes robiniophila Murrill, and its average molecular weight was estimated to be 8.7 × 10(4) Da. Monosaccharide composition analysis by gas chromatography (GC) indicated that TRP was composed of glucose, galactose, and arabinose in the molar ratio of 4.2:1.10:1.06. Particularly, we evaluated the anti-cancer efficacy of TRP on human osteosarcoma U-2 OS cells in vitro and associated possible molecular mechanism. Our result provided the first evidence that treatment of U-2 OS cells with TRP resulted in a dose- and time-dependent inhibitory effect on cell proliferation of U-2 OS cells and caused apoptotic death. Moreover, TRP induced the apoptosis of U-2 OS cells via a mitochondria-dependent pathway, as evidenced by an increase in Bax/Bcl-2 ratio, a loss of mitochondrial membrane potential (Δψm), release of cytochrome c from the mitochondria to the cytosol, activation of caspase-9 and caspase-3, and cleavage of poly(ADP-ribose) polymerase (PARP) in U-2 OS cells. In addition, overexpression of metadherin (MTDH), one carcinogene, was inhibited in U-2 OS cells after exposure to TRP for 24 h. Our findings suggested that TRP inhibited the proliferation of human osteosarcoma cancer cells by promoting apoptosis through the intrinsic mitochondrial pathway, as well as inhibition of MTDH expression.