Peripheral T cell receptor beta immune repertoire is promptly reconstituted after acute myocardial infarction

Abstract Background Acute myocardial infarction (AMI) is characterized by an inflammatory process in which T cell plays a key role. However, the profile of immune microenvironment in AMI is still uncertain. High-throughput sequencing of T cell receptor (TCR) provides deep insight into monitoring the immune microenvironment. Methods 30 patients with AMI were enrolled and 30 healthy individuals were recruited as controls. Flow cytometer were used to analyze the distribution of αβ T cells and their CD69 expression from peripheral leukomonocytes. TCRβ repertoire library was amplified by two-round multiplex PCR and detected by next-generation sequencing (NGS). Results The percentage of αβ T cells in AMI patients were significantly restricted than those in healthy controls, while the highly activated αβ T cells along with distinguishing usage of variable (V), diversity (D) and joining (J) gene segments were also found in AMI patients. In addition, AMI induced a significantly restricted CDR3 amino acid (AA) diversity and remarkably reconstituted TCR immune repertoires. Finally, we identified several AMI-associated tendency of CDR3 AAs expression after AMI. Conclusions Our work suggests that the aberrant αβ T cells distribution and activation may associated with the pathogenesis of AMI and demonstrates a reconstitution of TCRβ immune repertoire after AMI

Vancomycin-Stabilized Platinum Nanoparticles with Oxidase-like Activity for Sensitive Dopamine Detection

The development of efficient, reliable, and sensitive dopamine detection methods has attracted much attention. In this paper, vancomycin-stabilized platinum nanoparticles (Van-Ptn NPs, n = 0.5, 1, 2) were prepared by the biological template method, where n represented the molar ratio of vancomycin to Pt. The results show that Van-Pt2 NPs had oxidase-like activity and peroxidase-like activity, and the mechanism was due to the generation of reactive oxygen 1O2 and OH. Van-Pt2 NPs exhibited good temperature stability, storage stability, and salt solution stability. Furthermore, Van-Pt2 NPs had almost no cytotoxicity to A549 cells. More importantly, the colorimetric detection of DA in human serum samples was performed based on the oxidase-like activity of Van-Pt2 NPs. The linear range of DA detection was 10–700 μM, and the detection limit was 0.854 μM. This study establishes a rapid and reliable method for the detection of dopamine and extends the application of biosynthetic nanoparticles in the field of biosensing

Biocompatible Platinum Nanoclusters Prepared Using Bitter Gourd Polysaccharide for Colorimetric Detection of Ascorbic Acid

Ascorbic acid is an organic compound with antioxidant properties that can protect the human body from the threat of free radicals. Therefore, it is important to detect the existence and measure the concentration of ascorbic acid to regulate its content in the human body. In this work, we prepared bitter gourd polysaccharide (BGP)-stabilized platinum nanoclusters (Pt-BGP NCs) by reacting BGP with K2PtCl4. Pt-BGP NCs and catalyzed the decomposition of H2O2 to generate •OH radicals, which could oxidize TMB to form oxidized TMB (oxTMB), indicating their peroxidase-like properties. The kinetics followed the Michaelis–Menten equation. Furthermore, the colorimetric detection of ascorbic acid using Pt-BGP NCs showed high selectivity and a low detection limit of 0.191 μM. The accuracy of real sample detection using Pt-BGP NCs was as high as 98.9%. More importantly, Pt-BGP NCs had high cell biocompatibility and extremely low hemolysis rate due to the component of BGP. In summary, the prepared Pt-BGP NCs with reductive activity and good biocompatibility have good application prospects in colorimetric detection of ascorbic acid

The role of mitochondrial fusion and fission in the process of cardiac oxidative stress

Summary. Mitochondria are the energy suppliers in the cell and undergo constant fusion and fission to meet metabolic demand during the cell life cycle. Well- balanced mitochondrial dynamics are extremely important and necessary for cell survival as well as for tissue homeostasis. Cardiomyocytes contain large numbers of mitochondria to satisfy the high energy demand. It has been established that deregulated processes of mitochondrial dynamics play a major role in myocardial cell death. Currently, cardiac mitochondrial cell death pathways attract great attention in the cell biology and regenerative medicine fields. Importantly, mitochondrial dynamics are tightly linked to oxidative stress-induced cardiac damage. This review summarizes molecular mechanisms of mitochondrial fusion and fission processes and their potential roles in myocardial cell death triggered by oxidative stress. Advances in understanding the effect of both normal and abnormal mitochondrial dynamics on heart protection will lead to significant improvement of therapeutic discoverie

Sinomenine Suppresses Osteoclast Formation and <i>Mycobacterium tuberculosis</i> H37Ra-Induced Bone Loss by Modulating RANKL Signaling Pathways

<div><p>Receptor activator of NF-κB ligand (RANKL) is essential for osteoclastogenesis. Targeting RANKL signaling pathways has been an encouraging strategy for treating lytic bone diseases such as osteoporosis and rheumatoid arthritis (RA). Sinomenine (SIN), derived from Chinese medicinal plant <i>Sinomenioum</i><i>acutum</i>, is an active compound to treat RA, but its effect on osteoclasts has been hitherto unknown. In the present study, SIN was found to ameliorate <i>M. tuberculosis</i> H37Ra (Mt)-induced bone loss in rats with a decreased serum level of TRACP5b and RANKL, and an increased level of osteoprotegerin (OPG). <i>In vitro</i> study also showed that SIN could inhibit RANKL-induced osteoclast formation and bone resorption. The osteoclastic specific marker genes induced by RANKL including c-Src, MMP-9, TRACP were inhibited by SIN in a dose dependent manner. Signal transduction studies showed that SIN could obviously reduce the expression of RANK adaptor molecule TRAF6 and down-regulate RANKL-induced NF-κB activation. It decreased the RANKL-induced p38, JNK posphorylation but not ERK1/2 posphorylation. SIN could also reduce RANKL-mediated calcium influx which is associated with TRAF6/c-Src complex. Finally, SIN suppressed RANKL induced AP-1 and NFAT transcription, as well as the gene expression of NFATc1 and AP-1 components (Fra-1, Fra-2, c-Fos). The protein expression of c-Fos and TRAF6 were also inhibited by SIN after RANKL stimulation. Taken together, SIN could attenuate osteoclast formation and Mt-induced bone loss by mediating RANKL signaling pathways.</p> </div

Sinomenine ameliorated Mt-induced arthritis and bone loss in rats.

<p>SD rat was injected with 2 mg heat-killed M. tuberculosis H37Ra (Mt) in 200 µl mineral oil subcutaneously at the base of the tail at day 0 to induce arthritis (n=8). SIN was administrated intraperitoneally at a dose of 80 mg/kg/day. Another anti-rheumatic drug, TWP, was administrated orally at a dose of 2.5 mg/kg/day. Rats in control group were not immunized with Mt. (A) Arthritic edema was indicated by measuring the swelling volume of the hind paw. At day 28, rats were killed and the hind lambs were stocked in 10% formalin to obtain (B) the 2-D image of hind lamb and (C) the bone parameters by Micro-CT. (D) Serum prepared from rat abdominal aorta blood was used to measure RANKL, OPG, RANKL/OPG ratio, TRACP5b (showing the osteoclast activity), ALP activity (showing the osteoblast activity). All the bars are mean±SEM of a representative experiment. *P<0.05; **P<0.01 (compared with model group); # P<0.05, # #P<0.01 (compared with control group).</p

Sinomenine down-regulated RANKL-induced

<div><p>NF-κB <b>activation</b>. </p> <p>RAW264.7 cells, stably or transiently expressing NF-κB-luc reporter gene construct, were stimulated with RANKL for 8 h to study the effects of SIN on (A) stable or (B) transient NF-κB transcription. In another experiment, RAW264.7 cells were pretreated with SIN for 30 min and stimulated with 100 ng/ml RANKL for 30 min to (C) perform Western blot analysis of nuclear p65 and IκBα or (D) to observe p65 translocation to nuclear by fluorescence image. (C) The translocation of p65 by Western blot was expressed by dividing the nuclear p65 density with cytoplamic p65 density, the fold change of IκBα was normalized to β-actin. Densitometric quantification and statistical analysis include the results from 3 independent experiments. *P<0.05; **P<0.01 (compared with RANKL group); # P<0.05, # #P<0.01 (compared with control group).</p></div

Sinomenine reduced intracellular calcium influx.

<p>RAW264.7 cells were incubated with RANKL (100 ng/ml) in the presence or absence of SIN (0.5 mM) for 72 h. For Ca²⁺ measurement, cells were incubated with Fluo-3/AM for 30 min in serum-free DMEM followed by confocal analysis. (A) Intracellular calcium was illustrated by green fluorescence (Fluo-3/AM). (B) Each line represented the fluorescence intensity of one cell. The fluorescence intensity in each group was recorded within 200s and (C) statistically analyzed at time point of 200s. P<0.05; **P<0.01 (compared with RANKL group); # P<0.05, # #P<0.01 (compared with control group).</p

Sinomenine dose-dependently inhibited RANKL-induced osteoclast-specific genes and TRAF6.

<p>RAW264.7 cells were plated into 6-well plates, incubated with RANKL (100 ng/mL) and SIN for 24 h. Total RNA was extracted and real-time PCR was performed. Relative Quantification (RQ) was used to determine the fold-change of gene expression compared with the GAPDH control gene. (A) osteoclast-specific genes, (B) RANK and TRAF6 genes expression were investigated. RAW264.7 cells were incubated with SIN for 30 min and then stimulated with RANKL for 30 min, (C) TRAF6 protein expression was analyzed by Western blot. β-actin was used as loading control. Densitometric quantification and statistical analysis include the results from 3 independent experiments. *P<0.05; **P<0.01(compared with RANKL group); # P<0.05, # #P<0.01 (compared with control group).</p

Sinomenine inhibited RANKL-induced NFAT, AP-1 activation.

<p>RAW264.7 cells, transiently expressing NFAT-luc or AP-1-luc reporter gene construct were pre-treated with SIN for 30 min and followed by 100 ng/ml RANKL stimulation for 12 h. (A) NFAT and (B) AP-1 transcriptions are indicated by luciferase activity. (C) RAW264.7 cells were plated into 6-well plates, incubated with RANKL (100 ng/mL) and SIN for 24 h. Real-time PCR was performed to determine gene expressions of NFATc1 and AP-1 components (c-Fos, Fra-1, Fra-2). RAW264.7 cells were incubated with SIN for 30 min and then stimulated with RANKL for 30 min. (D) c-Fos protein expression was analyzed by Western blot. β-actin was used as loading control. Densitometric quantification and statistical analysis include the results from 3 independent experiments. P<0.05; **P<0.01(compared with RANKL group); # P<0.05, # #P<0.01 (compared with control group). (E) A schematic diagram shows a hypothetical model by which SIN inhibits osteoclast differentiation and function.</p