Tumour necrosis factor α regulates the miR-27a-3p-Sfrp1 axis in a mouse model of osteoporosis.

Osteoporosis is a metabolic bone disease that involves gradual loss of bone density and mass, thus resulting in increased fragility and risk of fracture. Inflammatory cytokines, such as tumour necrosis factor α (TNF-α), inhibit osteogenic differentiation of bone marrow mesenchymal stem cells (BMSCs), and several microRNAs are implicated in osteoporosis development. This study aimed to explore the correlation between TNF-α treatment and miR-27a-3p expression in BMSC osteogenesis and further understand their roles in osteoporosis. An osteoporosis animal model was established using ovariectomized (OVX) mice. Compared with Sham mice, the OVX mice had a significantly elevated level of serum TNF-α and decreased level of bone miR-27a-3p, and in vitro TNF-α treatment inhibited miR-27a-3p expression in BMSCs. In addition, miR-27a-3p promoted osteogenic differentiation of mouse BMSCs in vitro, as evidenced by alkaline phosphatase staining and Alizarin Red-S staining, as well as enhanced expression of the osteogenic markers Runx2 and Osterix. Subsequent bioinformatics analysis combined with experimental validation identified secreted frizzled-related protein 1 (Sfrp1) as a downstream target of miR-27a-3p. Sfrp1 overexpression significantly inhibited the osteogenic differentiation of BMSCs in vitro and additional TNF-α treatment augmented this inhibition. Moreover, Sfrp1 overexpression abrogated the promotive effect of miR-27a-3p on the osteogenic differentiation of BMSCs. Furthermore, the miR-27a-3p-Sfrp1 axis was found to exert its regulatory function in BMSC osteogenic differentiation via regulating Wnt3a-ß-catenin signalling. In summary, this study revealed that TNF-α regulated a novel miR-27a-3p-Sfrp1 axis in osteogenic differentiation of BMSCs. The data provide new insights into the development of novel therapeutic strategies for osteoporosis.

Multi-stage nuclear transcriptomic insights of morphogenesis and biparental role changes in Lentinula edodes.

Based on six offspring with different mitochondrial (M) and parental nuclear (N) genotypes, the multi-stage morphological characteristics and nuclear transcriptomes of Lentinula edodes were compared to investigate morphogenesis mechanisms during cultivation, the key reason for cultivar resistance to genotype changes, and regulation related to biparental role changes. Six offspring had specific transcriptomic data and morphological characteristics that were mainly regulated by the two parental nuclei, followed by the cytoplasm, at different growth stages. Importing a wild N genotype easily leads to failure or instability of fruiting; however, importing wild M genotypes may improve cultivars. Major facilitator superfamily (MFS) transporter genes encoding specific metabolites in spawns may play crucial roles in fruiting body formation. Pellets from submerged cultivation and spawns from sawdust substrate cultivation showed different carbon metabolic pathways, especially in secondary metabolism, degradation of lignin, cellulose and hemicellulose, and plasma membrane transport (mainly MFS). When the stage of small young pileus (SYP) was formed on the surface of the bag, the spawns inside were mainly involved in nutrient accumulation. Just broken pileus (JBP) showed a different expression of plasma membrane transporter genes related to intracellular material transport compared to SYP and showed different ribosomal proteins and cytochrome P450 functioning in protein biosynthesis and metabolism than near spreading pileus (NSP). Biparental roles mainly regulate offspring metabolism, growth, and morphogenesis by differentially expressing specific genes during different vegetative growth stages. Additionally, some genes encoding glycine-rich RNA-binding proteins, F-box, and folliculin-interacting protein repeat-containing proteins may be related to multi-stage morphogenesis. KEY POINTS: • Replacement of nuclear genotype is not suitable for cultivar breeding of L. edodes. • Some genes show a biparental role-divergent expression at mycelial growth stage. • Transcriptomic changes of some sawdust substrate cultivation stages have been elucidated.

Sappanone A Alleviated IL-1ß-Induced Inflammation in OA Chondrocytes through Modulating the NF-κB and Nrf2/HO-1 Pathways.

Objective: This study was to examine the anti-inflammatory effect of sappanone A on interleukin- (IL-) 1ß-stimulated osteoarthritis (OA) chondrocytes. Methods: Chondrocytes were pretreated with sappanone A for 2 h before subsequent IL-1ß stimulation. The mRNA expression levels of iNOs, COX-2, aggrecan, and collagen-II were measured with qRT-PCR. The levels of TNF-α, IL-6, IL-8, MMP-3, and MMP-13 were determined by ELISA. The protein levels of iNOs, COX-2, ADAMTS-4, ADAMTS-5, aggrecan, collagen-II, p-p65, p65, IκBα, Nrf2, and HO-1 were assessed by Western blot. Results: Sappanone A inhibited the IL-1ß-stimulated production of NO, PGE2, iNOS, COX-2, TNF-α, IL-6, and IL-8 in OA chondrocytes. In addition, sappanone A suppressed the expression of MMP-3, MMP-13, ADAMTS-4, and ADAMTS-5 in IL-1ß-stimulated OA chondrocytes. The degradation of ECM components was reversed by sappanone A. Sappanone A prevented NF-κB activation while enhanced Nrf2/HO-1 activation in IL-1ß-treated chondrocytes. Conclusion: Sappanone A may be a potent therapeutic agent for OA.

Enzyme-Powered Micro/Nanomotors for Cancer Treatment.

The incidence and lethal rate of cancers are rapidly rising recently, however current treatments of cancers, such as surgical resection, radiotherapy, chemotherapy and targeted therapy, usually require long treatment period and have more side effects and high recurrence rate. Enzyme-powered micro/nanomotors (EMNMs), with powerful self-propulsion, enhanced permeability and good biocompatibility, have shown great potential in crossing biological barrier and targeted drug transportation for cancer treatment; moreover, advanced approaches based on EMNMs such as photothermal therapy and starvation therapy have also been widely explored in cancer treatment. Although there are several review works discussing the progress of micro/nanomotors for biomedical applications, there is not one review paper with the focus on the cancer treatment based on EMNMs. Therefore, in this review, we try to concisely and timely summarize the recent progress of cancer treatment based on enzyme-driven micro/nanomotors, such as brain tumors, bladder cancer, breast cancer and others. Finally, the challenges and outlook of cancer therapy based on EMNMs are discussed, hoping to provide fundamental guidance for the future development.

Undercorrection: the undesired effect of compression on the osteotomy gap of the medial opening wedge high tibial osteotomy and its clinical significance.

INTRODUCTION: Undercorrection is a common problem in opening wedge high tibial osteotomy (OWHTO). We investigated the compression effect of cortex screw on the osteotomy gap and its clinical significance. MATERIALS AND METHODS: A standard OWHTO using the TomoFix plate was conducted on 20 bone models in two groups to get a 10-mm medial osteotomy gap. A cortex screw was used temporarily in a neutral (at the center) and an eccentric position (near the inclined plane) of the dynamic hole in group 1 and group 2, respectively. The mean of undercorrection observed in the two groups was compared using an independent t test. Also, the effect of compression on the gap between the plate and medial tibial cortex, and the osteotomy gap was evaluated using a Sine rule. Besides, the mean undercorrection observed was assessed for clinical significance based on the effect on the weight-bearing axis (WBA) using a Cosine Rule. RESULTS: The mean undercorrection was 1.3 ± 0.6 mm and 2.6 ± 0.6 mm in group 1 and group 2, respectively. A significantly greater undercorrection was observed in group 2 (p < 0.001). The correction loss in group 2 has resulted from combinations of the sliding effect of the dynamic hole and oblique compression effect over the gap between the plate and medial tibial cortex whereas in group 1 it has only resulted from the oblique compression effect. The observed undercorrection in group 2 has resulted in clinically significant WBA shift (10%) over the width of the tibial plateau. CONCLUSIONS: In OWHTO, compression is important for the stability and healing of osteotomy, but it can also cause loss of correction. In patients requiring large correction, the surgeon should control the amount of compression required and consider making extra osteotomy gap to avoid undercorrection. Furthermore, the placement of cortex screws in neutral is essential to lower the risk of undercorrection.

Preoperative Planning Using 3D Printing Technology in Orthopedic Surgery.

The applications of 3D printing technology in health care, particularly orthopedics, continue to broaden as the technology becomes more advanced, accessible, and affordable worldwide. 3D printed models of computed tomography (CT) and magnetic resonance image (MRI) scans can reproduce a replica of anatomical parts that enable surgeons to get a detailed understanding of the underlying anatomy that he/she experiences intraoperatively. The 3D printed anatomic models are particularly useful for preoperative planning, simulation of complex orthopedic procedures, development of patient-specific instruments, and implants that can be used intraoperatively. This paper reviews the role of 3D printing technology in orthopedic surgery, specifically focusing on the role it plays in assisting surgeons to have a better preoperative evaluation and surgical planning.

[Effects of different salinity levels on nitrification processes in sediments of Minjiang River Estuary, China]. / 不同盐度对闽江河口沉积物硝化作用的影响.

Two estuary wetlands in Minjiang River, Shanyutan and Daoqingzhou, were selected as the research objects. Wetland sediments were collected to examine the effects of different salinity levels on nitrification processes in the wetland with a culture experiment. The results showed that the nitrification rate of sediment in Minjiang River estuary wetland was generally low. The highest nitrification rate of sediment in the Shanyutan wetland was 0.193 mg·kg-1·d-1, while that in the Daoqingzhou wetland did not exceed 0.050 mg·kg-1·d-1. In the low salinity level (5), the decrease of nitrification rate was attributed to the restraint of nitrifying bacteria activities. The nitrification rate slightly increased with the increases of salinity (10), but was still lower than the initial value. This would be owed to the fact that the inhibitory effect of salinity on the activities of aerobic bacteria was strengthened, which reduced the rate of NH4+-N production, and thus resulted in a decrease of the contribution of aerobic ammonification bacteria to the apparent nitrification rate. There were regional variations in the responses of sediment nitrifying activity to salinity. In saltwater wetland (Shanyutan wetland), the adaptability of microbes in sediment to salinity was stronger, so that the nitrification activity in high salinity conditions was still higher. While in the freshwater wetland (Daoqingzhou wetland), the adaptability of the sediment to the salinity change was relatively lower, with a lower nitrification activity of the sediment in high salinity than in middle salinity. Acidic condition was the main reason for the low nitrification in the sediments of Minjiang estuary wetland. The nitrification rate and nitrification activity of the two wetlands increased first and then decreased with the duration of the culture experiment, which was driven by combined effects of initial NH4+-N concentration, oxygen content and denitrifying.

[Expression Level and Clinical Significance of VEGF, IL-17, ß2-MG and IL-35 in Patients with Multiple Myeloma].

OBJECTIVE: To explore the expression and clinical significance of VEGF, IL-17, ß2-MG and IL-35 in patients with multiple myeloma. METHODS: A total of 83 patients with multiple myeloma (MM) from January 2012 to December 2016 were enrolled in MM group, 36 healthy subjects were enrolled in control group. The levels of IL-17, IL-35 and VEGF in serum were detected by ELISA. The levels of ß2-MG in serum were measured by radioimmunoassay. The differences of different indexes between 2 groups were compared. RESULTS: The serum levels of IL-17, VEGF and ß2-MG in serum of III stage were higher than that in II stage, which was higher than that in I stage and control group (P<0.05). The levels of IL-35 in the control group were significantly higher than those in the I,II,III stage group (P<0.05). The levels of IL-17, VEGF and ß2-MG in serum of progress period were higher than those in stable phase and control group, level of IL-35 in serum of control group was significantly higher than that in the stable phase and progress period group (P<0.05). The correlation analysis showed that the level of serum IL-17 positively correlated with VEGF, ß2-MG expression (r=0.65, 0.58, P<0.05); and the serum IL-17 levels were negatively correlated with IL-35 levels (r=-0.42, P<0.05). CONCLUSION: The anomalous levels of IL-17, IL-35, IVEGF and ß2-MG expressions correlate with the progression and prognosis of patients with multiple myeloma.

High expression of TRIM29 (ATDC) contributes to poor prognosis and tumor metastasis by inducing epithelial­mesenchymal transition in osteosarcoma.

The association of TRIM29 overexpression with cancer progression and poor clinical prognosis has been reported in the context of several types of cancers. In the present study, we investigated the prognostic relevance of TRIM29 and its involvement in the progression of human osteosarcoma. To the best of our knowledge, this is the first study to demonstrate a major role of TRIM29 in osteosarcoma. Our results showed that the expression of TRIM29 in osteosarcoma tissues was much higher than that in normal bone tissues. Furthermore, TRIM29 expression was significantly correlated with tumor size, recurrence, metastasis and overall survival time. High expression of TRIM29 and presence of metastasis were independent predictors of poor prognosis in these patients. Both protein and mRNA expression of TRIM29 in osteosarcoma cell lines were significantly higher than those in osteoblast cell line, hFOB1.19. Moreover, the results indicated that TRIM29 promoted migration and invasive growth of osteosarcoma cells by inducing epithelial-mesenchymal transition. Therefore, ectopic expression of TRIM29 potentially contributes to metastasis and poor prognosis in patients with osteosarcoma. In summary, TRIM29 is a potential prognostic biomarker and a therapeutic target for patients with osteosarcoma.

Antidiabetic, antihyperlipidemic and antioxidant activities of a novel proteoglycan from ganoderma lucidum fruiting bodies on db/db mice and the possible mechanism.

Previously, we screened a proteoglycan for anti-hyperglycemic, named FYGL, from Ganoderma Lucidum. For further research of the antidiabetic mechanisms of FYGL in vivo, the glucose homeostasis, activities of insulin-sensitive enzymes, glucose transporter expression and pancreatic function were analyzed using db/db mice as diabetic models in the present work. FYGL not only lead to a reduction in glycated hemoglobin level, but also an increase in insulin and C-peptide level, whereas a decrease in glucagons level and showed a potential for the remediation of pancreatic islets. FYGL also increased the glucokinase activities, and simultaneously lowered the phosphoenol pyruvate carboxykinase activities, accompanied by a reduction in the expression of hepatic glucose transporter protein 2, while the expression of adipose and skeletal glucose transporter protein 4 was increased. Moreover, the antioxidant enzyme activities were also increased by FYGL treatment. Thus, FYGL was an effective antidiabetic agent by enhancing insulin secretion and decreasing hepatic glucose output along with increase of adipose and skeletal muscle glucose disposal in the late stage of diabetes. Furthermore, FYGL is beneficial against oxidative stress, thereby being helpful in preventing the diabetic complications.

[Effect of different perioperative treatments on gut flora in SD rats].

OBJECTIVE: To study the effects of different perioperative treatments on the number and proportion of gut flora in SD rats. METHODS: Forty-eight SD rats were randomized into 8 groups including the control group, antibiotics group, bowel preparation group, fasting group, antibiotic-bowel preparation group, antibiotics-bowel preparation-fasting group, bowel preparation-surgery-antibiotics-early postoperative feeding group (early feeding group), and bowel preparation-surgery-antibiotics-postoperative fasting group. The rats were sacrificed and stool specimens were collected from the cecum. Stools were diluted and transferred to selective medium. Bacteria counts were calculated after 48 hours of culture under constant temperature. The changes in gut flora between the different groups were compared in terms of E.coli, Bacteroides, Bifidobacterium, and Enterococcus. RESULTS: Compared with the control group, the total bacteria, Bacteroid, Enterococcus, Bifidobacterium were all significantly decreased(P<0.05), while the E.coli count and the bacillus/coccus ratio were significantly elevated(P<0.05). In the bowel preparation group, the total bacteria count, Bacteroid, Enterococcus, Bifidobacterium were all significantly decreased(P<0.05), while the E.coli count remained stable(P>0.05) and the bacillus/coccus ratio was significantly elevated(P<0.05). In the fasting group, the total bacteria count, Bacteroid, Enterococcus, Bifidobacterium were all significantly decreased(P<0.05), while the E.coli count remained stable(P>0.05) and the bacillus/coccus ratio was significantly elevated(P<0.05). Early postoperative feeding increased E.coli, Enterococcus, and total bacteria count(P<0.05), and lowered bacillus/coccus ratio(P<0.05) as compared to the fasting group. CONCLUSIONS: Antibiotics, bowel preparation, and fasting have influence on the gut flora of SD rats in count and bacillus/coccus ratio, leading to dysbiosis. Early postoperative feeding may improve dysbiosis.

Carrot antifreeze protein does not exhibit the polygalacturonase-inhibiting activity of PGIP family.

The carrot (Daucus carota) antifreeze protein (DcAFP) has a strong antifreeze activity and identified as belonging to the plant polygalacturonase-inhibiting protein (PGIP) family based on its sequence similarities, including the presence of a leucine-rich repeat (LRR) motif. In this study, yeast two-hybrid technology was used to analyze whether the carrot AFP could act as a PGIP. The complete DcAFP and polygalacturonase (PGase; obtained from fungus Alternaria alternata by RT-PCR) coding sequences were cloned into the bait and capture vectors, respectively, and yeast two-hybrid assays were performed. The results revealed that there was no evidence of an interaction between DcAFP and PGase, which suggests that DcAFP probably lacks PGIP activity. An analysis of the electrostatic potential of DcAFP and other PGIPs revealed that a large number of nonconservative residues within the beta-helix of the DcAFP LRR motif had been substituted to basic amino acids, thus changing the surface from negative to positive. This will electrostatically prevent DcAFP from binding with the positively charged surface of PGase. This is the first report that showed the correlation between nonconservative amino acids within the LRR motif of the DcAFP and its loss of polygalacturonase inhibiting activity.

Expression, purification, and antifreeze activity of carrot antifreeze protein and its mutants.

Antifreeze proteins (AFPs) enable organisms to survive under freezing or sub-freezing conditions. AFPs have a great potential in the low temperature storage of cells, tissues, organs, and foods. This process will require a large number of recombinant AFPs. In the present study, the recombinant carrot AFP was highly expressed in Escherichia coli strain BL21 (DE3). The activity of the purified and refolded recombinant proteins was analyzed by measurement of thermal hysteresis (TH) activity and detection of in vitro antifreeze activity by measuring enhanced cold resistance of bacteria. Two carrot AFP mutants generated by site-directed mutagenesis were also expressed and purified under these conditions for use in parallel experiments. Recombinant DcAFP displayed a TH activity equivalent to that of native DcAFP, while mutants DcAFP-N130Q and rDcAFP-N130V showed 32 and 43% decreases in TH activity, respectively. Both the recombinant DcAFP and its mutants were able to enhance the cold resistance of bacteria, to degrees consistent with their respective TH activities.

Significance of conservative asparagine residues in the thermal hysteresis activity of carrot antifreeze protein.

The approximately 24-amino-acid leucine-rich tandem repeat motif (PXXXXXLXXLXXLXLSXNXLXGXI) of carrot antifreeze protein comprises most of the processed protein and should contribute at least partly to the ice-binding site. Structural predictions using publicly available online sources indicated that the theoretical three-dimensional model of this plant protein includes a 10-loop beta-helix containing the approximately 24-amino-acid tandem repeat. This theoretical model indicated that conservative asparagine residues create putative ice-binding sites with surface complementarity to the 1010 prism plane of ice. We used site-specific mutagenesis to test the importance of these residues, and observed a distinct loss of thermal hysteresis activity when conservative asparagines were replaced with valine or glutamine, whereas a large increase in thermal hysteresis was observed when phenylalanine or threonine residues were replaced with asparagine, putatively resulting in the formation of an ice-binding site. These results confirmed that the ice-binding site of carrot antifreeze protein consists of conservative asparagine residues in each beta-loop. We also found that its thermal hysteresis activity is directly correlated with the length of its asparagine-rich binding site, and hence with the size of its ice-binding face.