TFAM loss induces nuclear actin assembly upon mDia2 malonylation to promote liver cancer metastasis.

The mechanisms underlying cancer metastasis remain poorly understood. Here, we report that TFAM deficiency rapidly and stably induced spontaneous lung metastasis in mice with liver cancer. Interestingly, unexpected polymerization of nuclear actin was observed in TFAM-knockdown HCC cells when cytoskeleton was examined. Polymerization of nuclear actin is causally linked to the high-metastatic ability of HCC cells by modulating chromatin accessibility and coordinating the expression of genes associated with extracellular matrix remodeling, angiogenesis, and cell migration. Mechanistically, TFAM deficiency blocked the TCA cycle and increased the intracellular malonyl-CoA levels. Malonylation of mDia2, which drives actin assembly, promotes its nuclear translocation. Importantly, inhibition of malonyl-CoA production or nuclear actin polymerization significantly impeded the spread of HCC cells in mice. Moreover, TFAM was significantly downregulated in metastatic HCC tissues and was associated with overall survival and time to tumor recurrence of HCC patients. Taken together, our study connects mitochondria to the metastasis of human cancer via uncovered mitochondria-to-nucleus retrograde signaling, indicating that TFAM may serve as an effective target to block HCC metastasis.

The effects of repeated freezing and thawing on bovine sperm morphometry and function.

Refreezing the remaining genetic resources after in vitro fertilization (IVF) can conserve genetic materials. However, the precise damage inflicted by repeated freezing and thawing on bovine sperm and its underlying mechanism remain largely unexplored. Thus, this study investigates the impact of repeated freeze-thaw cycles on sperm. Our findings indicate that such cycles significantly reduce sperm viability and motility. Furthermore, the integrity of the sperm plasma membrane and acrosome is compromised during this process, exacerbating the advanced apoptosis triggered by oxidative stress. Additionally, transmission electron microscopy exposed severe damage to the plasma membranes of both the sperm head and tail. Notably, the "9 + 2" structure of the tail was disrupted, along with a significant decrease in the level of the axonemal protein DNAH10, leading to reduced sperm motility. IVF outcomes revealed that repeated freeze-thaw cycles considerably impair sperm fertilization capability, ultimately reducing the blastocyst rate. In summary, our research demonstrates that repeated freeze-thaw cycles lead to a decline in sperm viability and motility, attributed to oxidative stress-induced apoptosis and DNAH10-related dynamic deficiency. As a result, the utility of semen is compromised after repeated freezing.

Syringin exerts anti-breast cancer effects through PI3K-AKT and EGFR-RAS-RAF pathways.

BACKGROUND: Breast cancer (BC) is one of the most common malignant tumors with the highest mortality in the world. Modern pharmacological studies have shown that Syringin has an inhibitory effect on many tumors, but its anti-BC efficacy and mechanism are still unclear. METHODS: First, Syringin was isolated from Acanthopanax senticosus (Rupr. & Maxim.) Harms (ASH) by systematic solvent extraction and silica gel chromatography column. The plant name is composed of genus epithet, species additive words and the persons' name who give its name. Then, the hub targets of Syringin against BC were revealed by bioinformatics. To provide a more experimental basis for later research, the hub genes which could be candidate biomarkers of BC and a ceRNA network related to them were obtained. And the potential mechanism of Syringin against BC was proved in vitro experiments. RESULTS: Syringin was obtained by liquid chromatography-mass spectrometry (LC-MS), nuclear magnetic resonance (NMR), and high-performance liquid chromatography (HPLC). Bioinformatics results showed that MAP2K1, PIK3CA, HRAS, EGFR, Caspase3, and PTGS2 were the hub targets of Syringin against BC. And PIK3CA and HRAS were related to the survival and prognosis of BC patients, the PIK3CA-hsa-mir-139-5p-LINC01278 and PIK3CA-hsa-mir-375 pathways might be closely related to the mechanism of Syringin against BC. In vitro experiments confirmed that Syringin inhibited the proliferation and migration and promoted apoptosis of BC cells through the above hub targets. CONCLUSIONS: Syringin against BC via PI3K-AKT-PTGS2 and EGFR-RAS-RAF-MEK-ERK pathways, and PIK3CA and HRAS are hub genes for adjuvant treatment of BC.

[Anti-herpes simplex virus type â of tectorigenin derivative and effect on Toll-like receptors in vitro].

The study investigated the inhibitory effect and mechanism of tectorigenin derivative(SGY) against herpes simplex virus type Ⅰ(HSV-1) by in vitro experiments. The cytotoxicity of SGY and positive drug acyclovir(ACV) on African green monkey kidney(Vero) cells and mouse microglia(BV-2) cells was detected by cell counting kit-8(CCK-8) method, and the maximum non-toxic concentration and median toxic concentration(TC_(50)) of the drugs were calculated. After Vero cells were infected with HSV-1, the virulence was determined by cytopathologic effects(CPE) to calculate viral titers. The inhibitory effect of the tested drugs on HSV-1-induced cytopathy in Vero cells was measured, and their modes of action were initially explored by virus adsorption, replication and inactivation. The effects of the drugs on viral load of BV-2 cells 24 h after HSV-1 infection and the Toll-like receptor(TLR) mRNA expression were detected by real-time fluorescence quantitative PCR(RT-qPCR). The maximum non-toxic concentrations of SGY against Vero and BV-2 cells were 382.804 µg·mL~(-1) and 251.78 µg·mL~(-1), respectively, and TC_(50) was 1 749.98 µg·mL~(-1) and 2 977.50 µg·mL~(-1), respectively. In Vero cell model, the half maximal inhibitory concentration(IC_(50)) of SGY against HSV-1 was 54.49 µg·mL~(-1), and the selection index(SI) was 32.12, with the mode of action of significantly inhibiting replication and directly inactivating HSV-1. RT-qPCR results showed that SGY markedly reduced the viral load in cells. The virus model group had significantly increased relative expression of TLR2, TLR3 and tumor necrosis factor receptor-associated factor 3(TRAF3) and reduced relative expression of TLR9 as compared with normal group, and after SGY intervention, the expression of TLR2, TLR3 and TRAF3 was decreased to different degrees and that of TLR9 was enhanced. The expression of inflammatory factors inducible nitric oxide synthase(iNOS), tumor necrosis factor-α(TNF-α), and interleukin-1ß(IL-1ß) was remarkably increased in virus model group as compared with that in normal group, and the levels of these inflammatory factors dropped after SGY intervention. In conclusion, SGY significantly inhibited and directly inactivated HSV-1 in vitro. In addition, it modulated the expression of TLR2, TLR3 and TLR9 related pathways, and suppressed the increase of inflammatory factor levels.

TFB2M activates aerobic glycolysis in hepatocellular carcinoma cells through the NAD+ /SIRT3/HIF-1α signaling.

BACKGROUND AND AIM: Increased aerobic glycolysis has been well-known as a hallmark of cancer, which is closely related to mitochondrial dysfunction. TFB2M (mitochondrial transcription factor B2) is a core mitochondrial transcription factor, which has been shown by us to play an oncogenic role in hepatocellular carcinoma (HCC). However, whether TFB2M contributes to the aerobic glycolysis in HCC cells remains unexplored. METHODS: The role and underlying molecular mechanisms of TFB2M in the regulation of aerobic glycolysis in HCC cells were systematically investigated by in vitro cell glucose metabolism and metabolomics analyses. Besides, the effects of TFB2M-regulated aerobic glycolysis in the growth and metastasis of HCC cells were also explored. RESULTS: Here, we show that TFB2M markedly enhanced the reprogramming of glucose metabolism from oxidative phosphorylation to aerobic glycolysis mainly through two mechanisms. On the one hand, TFB2M increased the expressions of glycolytic genes GAPDH, LDHA, GLUT1, and HK2. On the other hand, TFB2M decreased the expression of peroxisome proliferator-activated receptor gamma coactivator 1 alpha (PGC-1α), a critical regulator of mitochondrial respiration. Mechanistically, TFB2M regulates the upregulation of glycolytic genes and downregulation of PGC-1α mainly through NAD+ /SIRT3/HIF-1α signaling. Additionally, we found that TFBM2 promoted the progression of HCC cells through HIF-1α-regulated reprogramming of glucose metabolism. CONCLUSIONS: Our findings indicate that TFB2M serves as a critical glucose metabolic reprogramming mechanism in tumorigenesis, which could be used as potential therapeutic target in HCC.

Cardioprotective effect of rosmarinic acid against myocardial ischaemia/reperfusion injury via suppression of the NF-κB inflammatory signalling pathway and ROS production in mice.

CONTEXT: Rosmarinic acid (RosA), a natural poly-phenolic compound isolated from a variety of Labiatae herbs, has been reported to have a range of biological effects. OBJECTIVE: To investigate the cardioprotective effects of RosA against myocardial ischaemia/reperfusion (I/R) injury. MATERIALS AND METHODS: Male C57BL/6J mice were given RosA (100 mg/kg) via intragastric administration. After 1 week of administration, the mice were subjected to 30 min/24 h myocardial I/R injury. The mice were randomly subdivided into 4 groups: Vehicle, RosA, Vehicle + I/R, and RosA + I/R. Infarct size (IS), cardiac function (including EF, FS), histopathology, serum enzyme activities, ROS changes, cis aconitase (ACO) activity, and specific mRNA and protein levels were assessed in vivo. HL-1 cells were pre-treated with or without RosA (50 µM), followed by stimulation with 9 h/6 h of oxygen and glucose deprivation/re-oxygenation (OGD/R). The cells were randomly subdivided into 4 groups: Vehicle, RosA, Vehicle + OGD/R, and RosA + OGD/R. Lactate dehydrogenase (LDH) levels, ACO activity, ROS changes and protein levels were measured in vitro. RESULTS: Treatment with RosA reduced the following indicators in vivo (p < 0.05): (1) IS (14.5%); (2) EF (-23.4%) and FS (-18.4%); (3) the myocardial injury enzymes CK-MB (20.8 ng/mL) and cTnI (7.7 ng/mL); (4) DHE-ROS: (94.1%); (5) ACO activity (-2.1 mU/mg protein); (6) ogdh mRNA level (122.9%); and (7) OGDH protein level (69.9%). Moreover, treatment with RosA attenuated the following indicators in vitro (p < 0.05): (1) LDH level (191 U/L); (2) DHE-ROS: (165.2%); (3) ACO activity (-3.2 mU/mg protein); (4) ogdh mRNA level (70.0%); and (5) OGDH (110.1%), p-IκB-a (56.8%), and p-NF-κB (57.7%) protein levels. CONCLUSIONS: RosA has the potential to treat myocardial I/R injury with potential application in the clinic.

Flipping the cyclooxygenase (Ptgs) genes reveals isoform-specific compensatory functions.

Two prostaglandin (PG) H synthases encoded by Ptgs genes, colloquially known as cyclooxygenase (COX)-1 and COX-2, catalyze the formation of PG endoperoxide H2, the precursor of the major prostanoids. To address the functional interchangeability of these two isoforms and their distinct roles, we have generated COX-2>COX-1 mice whereby Ptgs2 is knocked in to the Ptgs1 locus. We then "flipped" Ptgs genes to successfully create the Reversa mouse strain, where knock-in COX-2 is expressed constitutively and knock-in COX-1 is lipopolysaccharide (LPS) inducible. In macrophages, flipping the two Ptgs genes has no obvious impact on COX protein subcellular localization. COX-1 was shown to compensate for PG synthesis at high concentrations of substrate, whereas elevated LPS-induced PG production was only observed for cells expressing endogenous COX-2. Differential tissue-specific patterns of expression of the knock-in proteins were evident. Thus, platelets from COX-2>COX-1 and Reversa mice failed to express knock-in COX-2 and, therefore, thromboxane (Tx) production in vitro and urinary Tx metabolite formation in COX-2>COX-1 and Reversa mice in vivo were substantially decreased relative to WT and COX-1>COX-2 mice. Manipulation of COXs revealed isoform-specific compensatory functions and variable degrees of interchangeability for PG biosynthesis in cells/tissues.

Fatty Acid Binding to the Allosteric Subunit of Cyclooxygenase-2 Relieves a Tonic Inhibition of the Catalytic Subunit.

Prostaglandin endoperoxide H synthase-2 (PGHS-2), also called cyclooxygenase-2 (COX-2), converts arachidonic acid to PGH2 PGHS-2 is a conformational heterodimer composed of allosteric (Eallo) and catalytic (Ecat) subunits. Fatty acids (FAs) bind to Arg-120 of Eallo increasing to different degrees, depending on the FA, the Vmax of its Ecat partner. We report here that movement of helical residues 120-122 and loop residues 123-129 of Eallo underlies the allosteric effects of FAs and allosteric COX-2 inhibitors, including naproxen and flurbiprofen. An S121P substitution in both PGHS-2 monomers yields a variant (S121P/S121P PGHS-2) that has 1.7-1.8 times the Vmax of native PGHS-2 and is relatively insensitive to activation by FAs or inhibition by allosteric inhibitors. The S121P substitution in Eallo is primarily responsible for these effects. In X-ray crystal structures, the Cα atoms of helical residues 119-122 of S121P/S121P PGHS-2 are displaced from their normal positions. Additionally, the S121P/S121P PGHS-2 variants in which Pro-127 and Ser-541 are replaced by cysteines spontaneously forms Cys-127 to Cys-541 cross-links between monomers. This is unlike the corresponding native PGHS-2 variant and suggests that S121P substitutions also unhinge the loop involving residues 123-129. We conclude the following: (a) the region involving residues 120-129 of unoccupied Eallo tonically inhibits Ecat; (b) binding of an activating FA (e.g. arachidonic, palmitic, or oleic acid) to Eallo or an S121P substitution in Eallo repositions this region to increase Ecat activity; and (c) allosteric COX inhibitors act by preventing FA binding to Eallo and additionally by relocating Eallo residues to inhibit Ecat.

Different Fatty Acids Compete with Arachidonic Acid for Binding to the Allosteric or Catalytic Subunits of Cyclooxygenases to Regulate Prostanoid Synthesis.

Prostaglandin endoperoxide H synthases (PGHSs), also called cyclooxygenases (COXs), convert arachidonic acid (AA) to PGH2. PGHS-1 and PGHS-2 are conformational heterodimers, each composed of an (Eallo) and a catalytic (Ecat) monomer. Previous studies suggested that the binding to Eallo of saturated or monounsaturated fatty acids (FAs) that are not COX substrates differentially regulate PGHS-1 versus PGHS-2. Here, we substantiate and expand this concept to include polyunsaturated FAs known to modulate COX activities. Non-substrate FAs like palmitic acid bind Eallo of PGHSs stimulating human (hu) PGHS-2 but inhibiting huPGHS-1. We find the maximal effects of non-substrate FAs on both huPGHSs occurring at the same physiologically relevant FA/AA ratio of ∼20. This inverse allosteric regulation likely underlies the ability of PGHS-2 to operate at low AA concentrations, when PGHS-1 is effectively latent. Unlike FAs tested previously, we observe that C-22 FAs, including ω-3 fish oil FAs, have higher affinities for Ecat than Eallo subunits of PGHSs. Curiously, C-20 ω-3 eicosapentaenoate preferentially binds Ecat of huPGHS-1 but Eallo of huPGHS-2. PGE2 production decreases 50% when fish oil consumption produces tissue EPA/AA ratios of ≥0.2. However, 50% inhibition of huPGHS-1 itself is only seen with ω-3 FA/AA ratios of ≥5.0. This suggests that fish oil-enriched diets disfavor AA oxygenation by altering the composition of the FA pool in which PGHS-1 functions. The distinctive binding specificities of PGHS subunits permit different combinations of non-esterified FAs, which can be manipulated dietarily, to regulate AA binding to Eallo and/or Ecat thereby controlling COX activities.

A Jurassic eutherian mammal and divergence of marsupials and placentals.

Placentals are the most abundant mammals that have diversified into every niche for vertebrates and dominated the world's terrestrial biotas in the Cenozoic. A critical event in mammalian history is the divergence of eutherians, the clade inclusive of all living placentals, from the metatherian-marsupial clade. Here we report the discovery of a new eutherian of 160 Myr from the Jurassic of China, which extends the first appearance of the eutherian-placental clade by about 35 Myr from the previous record, reducing and resolving a discrepancy between the previous fossil record and the molecular estimate for the placental-marsupial divergence. This mammal has scansorial forelimb features, and provides the ancestral condition for dental and other anatomical features of eutherians.

Interactions of 2-O-arachidonylglycerol ether and ibuprofen with the allosteric and catalytic subunits of human COX-2.

Prostaglandin (PG) endoperoxide H synthase (PGHS)-2, also known as cyclooxygenase (COX)-2, can convert arachidonic acid (AA) to PGH2 in the committed step of PG synthesis. PGHS-2 functions as a conformational heterodimer composed of an allosteric (Eallo) and a catalytic (Ecat) monomer. Here we investigated the interplay between human (hu)PGHS-2 and an alternative COX substrate, the endocannabinoid, 2-arachidonoylglycerol (2-AG), as well as a stable analog, 2-O-arachidonylglycerol ether (2-AG ether). We also compared the inhibition of huPGHS-2-mediated oxygenation of AA, 2-AG, and 2-AG ether by the well-known COX inhibitor, ibuprofen. When tested with huPGHS-2, 2-AG and 2-AG ether exhibit very similar kinetic parameters, responses to stimulation by FAs that are not COX substrates, and modes of inhibition by ibuprofen. The 2-AG ether binds Ecat more tightly than Eallo and, thus, can be used as a stable Ecat-specific substrate to examine certain Eallo-dependent responses. Ibuprofen binding to Eallo of huPGHS-2 completely blocks 2-AG or 2-AG ether oxygenation; however, inhibition by ibuprofen of huPGHS-2-mediated oxygenation of AA engages a combination of both allosteric and competitive mechanisms.

A cyclooxygenase-2-dependent prostaglandin E2 biosynthetic system in the Golgi apparatus.

Cyclooxygenases (COXs) catalyze the committed step in prostaglandin (PG) biosynthesis. COX-1 is constitutively expressed and stable, whereas COX-2 is inducible and short lived. COX-2 is degraded via endoplasmic reticulum (ER)-associated degradation (ERAD) following post-translational glycosylation of Asn-594. COX-1 and COX-2 are found in abundance on the luminal surfaces of the ER and inner membrane of the nuclear envelope. Using confocal immunocytofluorescence, we detected both COX-2 and microsomal PGE synthase-1 (mPGES-1) but not COX-1 in the Golgi apparatus. Inhibition of trafficking between the ER and Golgi retarded COX-2 ERAD. COX-2 has a C-terminal STEL sequence, which is an inefficient ER retention signal. Substituting this sequence with KDEL, a robust ER retention signal, concentrated COX-2 in the ER where it was stable and slowly glycosylated on Asn-594. Native COX-2 and a recombinant COX-2 having a Golgi targeting signal but not native COX-1 exhibited efficient catalytic coupling to mPGES-1. We conclude that N-glycosylation of Asn-594 of COX-2 occurs in the ER, leading to anterograde movement of COX-2 to the Golgi where the Asn-594-linked glycan is trimmed prior to retrograde COX-2 transport to the ER for ERAD. Having an inefficient ER retention signal leads to sluggish Golgi to ER transit of COX-2. This permits significant Golgi residence time during which COX-2 can function catalytically. Cytosolic phospholipase A2α, which mobilizes arachidonic acid for PG synthesis, preferentially translocates to the Golgi in response to physiologic Ca(2+) mobilization. We propose that cytosolic phospholipase A2α, COX-2, and mPGES-1 in the Golgi comprise a dedicated system for COX-2-dependent PGE2 biosynthesis.

[Antibacterial effect of self-etching adhesive systems on Streptococcus mutans].

OBJECTIVE: To investigate the antibacterial effect of different self-etching adhesive systems against Streptococcus mutans (S. mutans). METHODS: Six reagents Clearfil(TM) SE Bond primer (SP), Clearfil(TM) SE Bond adhesive (SA),Clearfil(TM) Protect Bond primer (PP), which contained antibacterial monomer methacryloyloxydodecylpyridinium bromide (MDPB), ClearfilTM Protect Bond adhesive (PA), positive control chlorhexidine acetate [CHX, 1% (mass fraction)], and negative control phosphate buffer solution (PBS) were selected. They were mixed with S. mutans for 30 s respectively, then colony-forming units (CFU) were counted after incubated for 48 h on brain heart infusion (BHI) agar medium. The 6 reagents were applied to the sterile paper discs, and distributed onto the BHI agar medium with S. mutans and incubated for 24 h, then the inhibition zones were observed. CHX, PBS, PP, and SP were added on the dentin with artificial caries induced by S. mutans and kept for 30 s, then confocal laser scanning microscope (CLSM) was used to observe the live and dead bacteria after staining. The ratio of live to dead bacteria was calculated. PP+PA and SP+SA were applied on the dentin according to the manual and light cured. S. mutans were incubated on the samples for 2 h, ultrasonically treated and incubated on BHI agar medium for 48 h, then CFU was counted. The data were analyzed by non-parametric analysis and one-way ANOVA. RESULTS: Compared with PBS, the PP, SP, PA, SA and CHX showed the antibacterial effect on free S. mutans (P<0.05); SP and PP showed stronger antibacterial effect than PA, SA and CHX (P<0.05). CHX, SP and PP presented inhibition zones, while PBS, SA and PA did not. Compared with PBS, the CHX, SP and PP could lower the ratio of the live to dead bacteria significantly (P<0.05). Cured self-etching adhesive systems did not show any antibacterial effect on the free S. mutans. CONCLUSION: The primer of self-etching adhesives Clearfil(TM) SE Bond and Clearfil(TM) Protect Bond showed significant antibacterial effect on free and attached S. mutans. The adhesive only showed antibacterial effect on free S. mutans before light-cured polymerization. After being cured, the self-etching adhesive systems did not show antibacterial effect anymore.

Azoxystrobin, a mitochondrial complex III Qo site inhibitor, exerts beneficial metabolic effects in vivo and in vitro.

BACKGROUND: Several anti-diabetes drugs exert beneficial effects against metabolic syndrome by inhibiting mitochondrial function. Although much progress has been made toward understanding the role of mitochondrial function inhibitors in treating metabolic diseases, the potential effects of these inhibitors on mitochondrial respiratory chain complex III remain unclear. METHODS: We investigated the metabolic effects of azoxystrobin (AZOX), a Qo inhibitor of complex III, in a high-fat diet-fed mouse model with insulin resistance in order to elucidate the mechanism by which AZOX improves glucose and lipid metabolism at the metabolic cellular level. RESULTS: Acute administration of AZOX in mice increased the respiratory exchange ratio. Chronic treatment with AZOX reduced body weight and significantly improved glucose tolerance and insulin sensitivity in high-fat diet-fed mice. AZOX treatment resulted in decreased triacylglycerol accumulation and down-regulated the expression of genes involved in liver lipogenesis. AZOX increased glucose uptake in L6 myotubes and 3T3-L1 adipocytes and inhibited de novo lipogenesis in HepG2 cells. The findings indicate that AZOX-mediated alterations to lipid and glucose metabolism may depend on AMP-activated protein kinase (AMPK) signaling. CONCLUSIONS: AZOX, a Qo inhibitor of mitochondrial respiratory complex III, exerts whole-body beneficial effects on the regulation of glucose and lipid homeostasis in high-fat diet-fed mice. GENERAL SIGNIFICANCE: These findings provide evidence that a Qo inhibitor of mitochondrial respiratory complex III could represent a novel approach for the treatment of obesity.

[Identification of Streptococcus mutans in carious patients' saliva samples by MALDI-TOF mass spectrometry].

OBJECTIVE: To identify Streptococcus mutans (S. mutans) in carious patients' saliva using matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS), and to establish a faster and more accurate method to identify S. mutans. METHODS: In this study, a total of 90 carious patients from Department of Endodontics of Peking University School of Stomatology were recruited. All these patients' saliva was collected. After extracting the protein of the samples, they were identified by MALDI-TOF MS. The composite profile was analyzed using BioExplorer 1.0 software. The scores ≥ 25 were considered as S. mutans, whereas the scores <25 were as considered as non S. mutans. Finally, these results were compared with 16S rDNA sequencing to figure out the sensitivity and concordance rate, respectively. RESULTS: The sensitivity of MALDI-TOF MS was 96.0%, and the concordance rate compared with 16S rDNA sequencing was as high as 98.7%. CONCLUSION: MALDI-TOF MS is high throughput, rapid and easy to perform in comparison to other conventional methods. It has a high sensivity and concordance rate. Thus, MALDI-TOF MS can serve as an effective tool for identification of S. mutans.

Electroacupuncture improves peripheral neuropathy by up-regulating Sirt1/PGC-1α/TFAM pathway in type 2 diabetes rats with peripheral neuropathy. / 基于Sirt1/PGC-1α/TFAM通路探讨电针治疗2åž‹ç³–å°¿ç— å‘¨å›´ç¥žç»ç— å˜çš„æœºåˆ¶.

OBJECTIVES: To observe the effect of electroacupuncture (EA) on activation of silent information regulator 1 (Sirt1)/peroxisome proliferator-activated receptor γ coactivator 1α (PGC-1α)/mitochondrial transcription factor A (TFAM) pathway in type 2 diabetes (T2DM) rats with peripheral neuropathy (DPN) , so as to explore its possible mechanisms underlying improvement of DPN. METHODS: Thirty male SD rats were randomly divided into blank control group (n=8) and DPN model group (n=22) which were further divided into model group (n=8) and EA group (n=8) after successful modeling. The model of T2DM was established by high-fat diet and low-dose intraperitoneal injection of streptozocin (35 mg/kg). For rats of the EA group (anesthetized with isoflurane), EA stimulation (2 Hz/15 Hz, 2 mA) was applied to "Tianshu"(ST25) for 20 min, once daily, 6 times a week for 6 weeks. The blood glucose level, body weight, area under curve (AUC) of glucose tolerance test, and hind-paw mechanical pain threshold and thermal pain threshold were observed. The intra-epidermal nerve fiber density (IENFD) of the hind-foot pad was observed by immunofluorescence staining. The motor nerve conduction velocity (MNCV) of the sciatic nerve was measured by using electrophysiological method. H.E. staining was used to observe the histopathological changes of the sciatic nerve after modeling. Transmission electron microscopy (TEM) was used to observe the ultrastructural changes of the sciatic nerve. The protein expressions of energy-related Sirt1, PGC-1α and TFAM in the sciatic nerve was detected by Western blot. RESULTS: Compared with the blank control group, the model group had a higher blood glucose contents and AUC (P<0.001), a slower MNCV (P<0.01), and a decrease in the body weight and in the mechanical and thermal pain thresholds (P<0.001) and IENFD (P<0.001), and in the expression levels of Sirt1, PGC-1α and TFAM (P<0.05, P<0.01). In contrast to the model group, the EA group had a decrease in the blood glucose contents and AUC (P<0.05, P<0.01), and an increase in mechanical and thermal pain thresholds, MNCV, IENFD, and expression levels of Sirt1, PGC-1α and TFAM proteins (P<0.01, P<0.05). In addition, results of histopathological and ultrastructural changes of the sciatic nerve showed more fragmented and disordered distribution of axons on the transverse section, and extensive separation of myelin and axons, uneven myelin thickness, axonal degeneration and irregular shape in the model group, whereas in the EA group, the axons on the transverse section were relatively more dense and more complete, the myelin sheath of the sciatic nerve was relatively uniform, and the axonal shape was relatively regular with relatively milder lesions. CONCLUSIONS: EA up-regulates the expressions of Sirt1, PGC-1α, TFAM in T2DM rats with DPN, which may be associated with its functions in improving and repairing the injured peripheral nerves in rats with DPN.

Assessment of isthmus filling using two obturation techniques performed by students with different levels of clinical experience.

Background/purpose: Root canal filling is a necessary skill for dental students and an important aspect of endodontic education. This study aimed to evaluate the effect of students' clinical experiences on isthmus filling using different techniques and sealers. Materials and methods: One hundred eight three-dimensional-printed resin replicas of isthmus were divided into six groups and either continuous wave of condensation (CWC) or single-cone obturation (SC) was performed. One of three sealers (AH Plus Jet®, GuttaFlow2, iRoot SP) was used together with a size-fitted gutta-percha master cone. All the obturations were completed by students with three different levels of clinical experience including senior postgraduate students (SPS), junior postgraduate students (JPS), and undergraduate students (US). The percentages of filled areas (PFA) at 2, 4, 6, and 8 mm from the apex were analyzed using a light microscope. Data were analyzed using the Mann-Whitney U test or Kruskal-Wallis 1-way ANOVA with Dunn's tests (α = 0.05). Results: The CWC group exhibited a higher PFA than the SC group (P < 0.05). The PFA was higher in the SPS group than in the JPS group or the US group with CWC (P < 0.05). The three clinical experience groups showed similar PFAs with SC (P > 0.05); however, when using SC with iRoot SP, the PFA was higher than with either of the other two sealers (P < 0.05). Conclusion: CWC was found to be technique-sensitive and required clinical training. With SC, clinical experience did not improve the quality of isthmus filling without additional training. CWC was superior to SC for type IV isthmuses. When using SC, better filling quality was obtained with a bioceramic sealer.

Polyurea-urethane Temperature-responsive Hydrogels for Sustained Delivery of Anti-VEGF Therapeutics.

Temperature-responsive hydrogels, or thermogels, have emerged as a leading platform for sustained delivery of both small molecule drugs and macromolecular biologic therapeutics. Although thermogel properties can be modulated by varying the polymer's hydrophilic-hydrophobic balance, molecular weight and degree of branching, varying the supramolecular donor-acceptor interactions on the polymer remains surprisingly overlooked. Herein, to study the influence of enhanced hydrogen bonding on thermogelation, we synthesized a family of amphiphilic polymers containing urea and urethane linkages using quinuclidine as an organocatalyst. Our findings showed that the presence of strongly hydrogen bonding urea linkages significantly enhanced polymer hydration in water, in turn affecting hierarchical polymer self-assembly and macroscopic gel properties such as sol-gel phase transition temperature and gel stiffness. Additionally, analysis of the sustained release profiles of Aflibercept, an FDA-approved protein biologic for anti-angiogenic treatment, showed that urea bonds on the thermogel were able to significantly alter the drug release mechanism and kinetics compared to usage of polyurethane gels of similar composition and molecular weight. Our findings demonstrate the unrealized possibility of modulating gel properties and outcomes of sustained drug delivery through judicious variation of hydrogen bonding motifs on the polymer structure.

Critical role of transmethylation in TLR signaling and systemic lupus erythematosus.

Post-translational protein modifications can play a significant role in immune cell signaling. Recently, we showed that inhibition of transmethylation curtails experimental autoimmune encephalomyelitis, notably by reducing T cell receptor (TCR)-induced activation of CD4(+) T cells. Here, we demonstrate that transmethylation inhibition by a reversible S-adenosyl-l-homocysteine hydrolase inhibitor (DZ2002) led to immunosuppression by reducing TLR-, B cell receptor (BCR)- and TCR-induced activation of immune cells, most likely by blocking NF-κB activity. Moreover, prophylactic treatment with DZ2002 prevented lupus-like disease from developing in both BXSB and MRL-Fas(lpr) mouse models. DZ2002 treatment initiated during active disease significantly improved outcomes in both in vivo models, suggesting methylation inhibition as a novel approach for the treatment of autoimmune/inflammatory diseases.