[Identification and expression analysis of whole gene family of Isatis indigotica 4-coumarate: CoA ligase].

The 4-coumarate: CoA ligase(4CL) is a key enzyme in the upstream pathway of phenylpropanoids such as flavonoids, soluble phenolic esters, lignans, and lignins in plants. In this study, 13 4CL family members of Arabidopsis thaliana were used as reference sequences to identify the 4CL gene family candidate members of Isatis indigotica from the reported I. indigotica genome. Further bioinformatics analysis and analysis of the expression pattern of 4CL genes and the accumulation pattern of flavonoids were carried out. Thirteen 4CL genes were obtained, named Ii4CL1-Ii4CL13, which were distributed on chromosomes 1, 2, 3, 4, and 6. The analysis of the gene structure and conserved structural domains revealed the intron number of I. indigotica 4CL genes was between 1 and 12 and the protein structural domains were highly conserved. Cis-acting element analysis showed that there were multiple response elements in the promoter sequence of I. indigotica 4CL gene family, and jasmonic acid had the largest number of reaction elements. The collinearity analysis showed that there was a close relationship between the 4CL gene family members of I. indigotica and A. thaliana. As revealed by qPCR results, the expression analysis of the 4CL gene family showed that 10 4CL genes had higher expression levels in the aboveground part of I. indigotica. The content assay of flavonoids in different parts of I. indigotica showed that flavonoids were mainly accumulated in the aboveground part of plants. This study provides a basis for further investigating the roles of the 4CL gene family involved in the biosynthesis of flavonoids in I. indigotica.

Downregulated calmodulin expression contributes to endothelial cell impairment in diabetes.

Endothelial dysfunction, a central hallmark of cardiovascular pathogenesis in diabetes mellitus, is characterized by impaired endothelial nitric oxide synthase (eNOS) and NO bioavailability. However, the underlying mechanisms remain unclear. Here in this study, we aimed to identify the role of calmodulin (CaM) in diabetic eNOS dysfunction. Human umbilical vein endothelial cells and murine endothelial progenitor cells (EPCs) treated with high glucose (HG) exhibited downregulated CaM mRNA/protein and vascular endothelial growth factor (VEGF) expression with impeded eNOS phosphorylation and cell migration/tube formation. These perturbations were reduplicated in CALM1-knockdown cells but prevented in CALM1-overexpressing cells. EPCs from type 2 diabetes animals behaved similarly to HG-treated normal EPCs, which could be rescued by CALM1-gene transduction. Consistently, diabetic animals displayed impaired eNOS phosphorylation, endothelium-dependent dilation, and CaM expression in the aorta, as well as deficient physical interaction of CaM and eNOS in the gastrocnemius. Local CALM1 gene delivery into a diabetic mouse ischemic hindlimb improved the blunted limb blood perfusion and gastrocnemius angiogenesis, and foot injuries. Diabetic patients showed insufficient foot microvascular autoregulation, eNOS phosphorylation, and NO production with downregulated CaM expression in the arterial endothelium, and abnormal CALM1 transcription in genome-wide sequencing analysis. Therefore, our findings demonstrated that downregulated CaM expression is responsible for endothelium dysfunction and angiogenesis impairment in diabetes, and provided a novel mechanism and target to protect against diabetic endothelial injury.

Wound healing can be improved by (-)-epigallocatechin gallate through targeting Notch in streptozotocin-induced diabetic mice.

Delayed wound healing is one of the most prominent clinical manifestations of diabetes and lacks satisfactory treatment options. Persistent inflammation occurs in the late phase of wound healing and impairs the healing process in mice with diabetes mellitus (DM). In this study, we observed that the late wound healing in streptozotocin (STZ)-induced DM mice could be improved by (-)-epigallocatechin gallate (EGCG). The macrophage accumulation, inflammation response, and Notch signaling can be inhibited by EGCG in the skin wounds of DM mice. Furthermore, we found that the LPS-induced inflammation response including overactivated Notch signaling, was inhibited by EGCG in mouse macrophages. Moreover, we confirmed that EGCG could directly bind with mouse Notch-1. In addition, our studies indicated that diabetic wound healing was improved by EGCG treatment before or after the inflammation phase by targeting the Notch signaling pathway, which suggests that the pre-existing diabetic wound healing can be improved by EGCG. To summarize, wound healing can be improved by EGCG through targeting Notch in STZ-induced DM mice. Our findings provide insight into the therapeutic strategy for diabetic wounds and offer EGCG as a novel potential medicine to treat chronic wounds.-Huang, Y.-W., Zhu, Q.-Q., Yang, X.-Y., Xu, H.-H., Sun, B., Wang, X.-J., Sheng, J. Wound healing can be improved by (-)-epigallocatechin gallate through targeting Notch in streptozotocin-induced diabetic mice.

Rhopaladins' analogue (E)-2-aroyl-4-(4-fluorobenzylidene)-5-oxopyrrolidines inhibit proliferation, promote apoptosis and down-regulation of E6/E7 mRNA in cervical cancer.

The occurrence and development of cervical cancer threaten women's life and health, HPV-induced cervical cancer is a major health issue among women. We synthesized three Rhopaladins' analogue (E)-2-aroyl-4-(4-fluorobenzylidene)-5-oxopyrrolidines via a tandem Ugi 4CC/SN cyclization with pyrrolidone as a core structure. In addition, the cytotoxicity of these new compounds in the cervical cancer cell line CaSki was studied by MTT assay. And then we chose one to research the apoptosis and the expression of E6/E7 mRNA in CaSki cells. The results indicated that the new compound can not only inhibited the proliferation of CaSki in dose-dependent and time-dependent manners but also induced the apoptosis, which may be related to the down-regulation of E6/E7 mRNA expression.

Antinociceptive roles of galanin receptor 1 in nucleus accumbens of rats in a model of neuropathic pain.

It has been reported that galanin and its receptors might be involved in the modulation and transmission of nociception in the central nervous system. Our previous research has also demonstrated that galanin induces antinociception in the nucleus accumbens (NAc) of intact rats. However, the interaction between galanin and its receptors in the NAc and the underlying mechanism of suppressing pain transmission remain unclear. The present study seeks to determine the antinociception induced by galanin receptor (GalR)-1 stimulation in the NAc of rats with neuropathic pain. The left sciatic nerve of rats was ligated to mimic a neuropathic pain model. Western blots showed that the expression of GalR1 was significantly upregulated in the NAc of rats with neuropathic pain. Intra-NAc injection of GalR1 agonist M617 induced a dose-dependent increase in hindpaw withdrawal latency (HWL) to noxious thermal and mechanical stimulations in rats with neuropathic pain. Also, the effect of M617 was attenuated by M35, a GalR1/2 antagonist; at the same time, M35 reduced the galanin-induced antinociception, suggesting that GalR1 mediates antinociception induced by galanin in the NAc of rats with neuropathic pain. Furthermore, we found that M617-induced antinociception in rats with neuropathic pain was stronger than the antinociception in intact rats. We also found that injections of M617 and galanin each induced significant increases in HWL, but the galanin-induced antinociception was stronger than that of M617. All these results suggest that GalR1 plays an important role in antinociception and that other GalRs also are involved in pain modulation induced by galanin in the NAc of rats with neuropathic pain.

Urolithin B Attenuates Cerebral Ischemia-reperfusion Injury by Modulating Nrf2-regulated Anti-oxidation in Rats.

Ischemia-reperfusion (IR) induces a wide range of irreversible injuries. Cerebral IR injury (IRI) refers to additional brain tissue damage that occurs after blood flow is restored following cerebral ischemia. Currently, no established methods exist for treating IRI. Oxidative stress is recognized as a primary mechanism initiating IRI and a crucial focal target for its treatment. Urolithin B, a metabolite derived from ellagitannins, antioxidant polyphenols, has demonstrated protective effects against oxidative stress in various disease conditions. However, the precise mechanism underlying UB's effect on IRI remains unclear. In our current investigation, we assessed UB's ability to mitigate neurological functional impairment induced by IR using a neurological deficit score. Additionally, we examined cerebral infarction following UB administration through TTC staining and neuron Nissl staining. UB's inhibition of neuronal apoptosis was demonstrated through the TUNEL assay and Caspase-3 measurement. Additionally, we examined UB's effect on oxidative stress levels by analyzing malondialdehyde (MDA) concentration, superoxide dismutase (SOD) activity, and immunohistochemistry analysis of inducible nitric oxide synthase (iNOS) and 8-hydroxyl-2'-deoxyguanosine (8-OHdG). Notably, UB demonstrated a reduction in oxidative stress levels. Mechanistically, UB was found to stimulate the Nrf2/HO-1 signaling pathway, as evidenced by the significant reduction in UB's neuroprotective effects upon administration of ATRA, an Nrf2 inhibitor. In summary, UB effectively inhibits oxidative stress induced by IR through the activation of the Nrf2/HO-1 signaling pathway. These findings suggest that UB holds promise as a therapeutic agent for the treatment of IRI.

Heterogeneous catalytic oxidation of glycerol over a UiO-66-derived ZrO2@C supported Au catalyst at room temperature.

The catalytic conversion of biomass-derived glycerol into high-value-added products, such as glyceric acid (GLYA), using catalyst-supported Au nanoparticles (Au NPs) at room temperature presents a significant challenge. In this study, we constructed a series of supported Au catalysts, including Au/ZrO2@C, Au/C, Au/ZrO2, and Au/ZrO2-C, and investigated their effectiveness in selectively catalytic oxidizing glycerol to GLYA at room temperature. Among these catalysts, the Au/ZrO2@C catalyst exhibited the best catalytic performance, achieving a glycerol conversion rate of 73% and a GLYA selectivity of 79% under the optimized reaction conditions (reaction conditions: 30 mL 0.1 M glycerol, glycerol/Au = 750 mol mol-1, T = 25 °C, p(O2) = 10 bar, stirring speed = 600 rpm, time = 6 h). Physical adsorption, X-ray diffraction (XRD), X-ray photoelectron spectroscopy (XPS), transmission electron microscopy (TEM), and other characterization methods were employed to analyze the texture properties of the catalyst. The findings indicated that the support structure, the strong metal-support interactions between Au NPs and the support, and the presence of small metallic Au NPs were the primary factors contributing to the catalyst's high activity and selectivity. Moreover, the reusability of the Au/ZrO2@C catalyst was investigated, and a probable reaction mechanism for the oxidation of glycerol was proposed.

Research Progress on Structure and Anti-Gynecological Malignant Tumor of Shikonin.

Gynecological malignancy seriously threatens the physical and mental health of women. Shikonin is a naphthoquinone compound with a variety of biological activities. Studies have shown that shikonin can inhibit cell proliferation, promote cell apoptosis and induce cell necrosis. And in recent years, shikonin are also being increasingly used for the study of gynecological malignant diseases. Therefore, we reviewed the mechanism of action and structure optimization of shikonin in gynecological malignant tumors, in order to provide some reference for further research and development of related drug.

Molecular cloning, identification and functional characterization of a novel intracellular Cu-Zn superoxide dismutase from the freshwater mussel Cristaria plicata.

Superoxide dismutases (SODs, EC 1.15.1.1) are one family of important antioxidant metalloenzymes involved in scavenging the high level of reactive oxygen species (ROS) into molecular oxygen and hydrogen peroxide. In the present study, the intracellular CuZnSOD gene of Cristaria plicata (Cp-icCuZnSOD) was identified from hemocytes by homology cloning and the rapid amplification of cDNA ends (RACE) technique. The full-length cDNA of Cp-icCuZnSOD consisted of 891 nucleotides with a canonical polyadenylation signal sequence ATTAAA, a poly (A) tail, and an open-reading frame of 468 bp encoding 155 amino acids. The deduced amino acids of CpSOD shared high similarity with the known icCuZnSODs from other species, and several highly conserved motifs including Cu/Zn ions binding sites (His-46, His-48, His-63, His-120 for Cu(2+) binding, and His-63, His-71, His-80, Asp-83 for Zn(2+) binding), intracellular disulfide bond and two CuZnSOD family signatures were also identified in CpSOD. Furthermore, the recombinant Cp-icCuZnSOD with high enzyme activity was induced to be expressed as a soluble form by IPTG supplemented with Cu/Zn ions at 20 degrees C for 8 h, and then was purified by using the native Ni(2+) affinity chromatography. The specific activity of the purified rCp-icCuZnSOD enzyme was 5368 U/mg, which is 2.6-fold higher than that of zebrafish Danio rerio rZSOD and 5.3-fold higher than that of bay scallop Argopecten irradians rAi-icCuZnSOD. The enzyme stability assay showed that the purified rCp-icCuZnSOD enzyme maintained more than 80% activity at temperature up to 60 degrees C, at pH 2.0-9.0, and was resistant to 8 mol/L urea or 8% SDS. In addition, the addition of active rCp-icCuZnSOD enzmye could protect hepatocyte L02 cells from oxidative damage as assessed using an alcohol-injured human liver cell model.

[Recovery of Phosphorus in the Form of Struvite from the Anaerobic Fermentation Supernatant of Excess Sludge].

To recover nitrogen and phosphorus from the supernatant of anaerobic fermentation of excess sludge in the form of struvite (MAP, MgNH4PO4·6H2O), the optimum conditions of nitrogen and phosphorus dissolution and the effects of different reaction conditions on nitrogen and phosphorus recovery were studied. The results showed that the concentration of ammonia nitrogen and orthophosphate that dissolved in the fermentation liquid peaked on the fifth day of the experiment at pH 10.5 and 35℃. When a phosphorus source was and was not added, the optimum conditions for recovering phosphorus were pH=9.5, N:P=0.8, and Mg:P=1.8 and pH=9.5, Mg:P=1.6, and speed=200 r·min-1, respectively. In addition, reducing the N:P molar ratio had a significant effect on the morphology and purity of the struvite. Scanning electron microscopy (SEM), energy dispersive spectrometry (EDS), X-ray diffraction (XRD), and Fourier transform infrared spectroscopy (FTIR) were used to analyze the surface appearance and phase composition of the recovered products. The main component of the precipitate was MAP. Recovery of nitrogen and phosphorus in excess sludge in the form of struvite is an effective means for realizing sludge resource utilization.

Absorption of caffeine in fermented Pu-er tea is inhibited in mice.

Caffeine is present in a number of dietary sources consumed worldwide. Although its pharmacokinetics has been intensively explored, little is known about complexed caffeine (C-CAF) in aqueous extraction of fermented Pu-er tea. The major components of C-CAF are oxidative tea polyphenols (OTP) and caffeine. Furthermore, the C-CAF can be precipitated in low pH solution. After administering the same amount of total caffeine and comparing the peak level of plasma caffeine with the coffee (contains 0.11 ± 0.01% C-CAF) group, the results showed that the caffeine/OTP (contains 66.67 ± 0.02% C-CAF) group and the instant Pu-er tea (contains 23.18 ± 0.02% C-CAF) group were 33.39% and 25.86% lower, respectively. The concentration of the metabolites of caffeine supports the idea that the absorption of the C-CAF was inhibited in mice. Congruent with this result, the amount of caffeine detected in mice excrement showed that more caffeine was eliminated in the caffeine/OTP group and the Pu-er tea group. The locomotor activity tests of mice demonstrated that the stimulating effect of caffeine in caffeine/OTP and Pu-er tea was weaker than in coffee. Our findings demonstrate that caffeine can be combined with OTP and the absorption of C-CAF is inhibited in mice, thus decreasing the irritation effect of caffeine. This may also be developed as a slow release formulation of caffeine.

Determination of trace mercury by solid substrate room temperature phosphorescence quenching method based on lead carboxymethyl cellulose (Pb(CMC)(2)) particles containing luminescent salicyl fluorones molecules.

Luminescent particles of lead carboxymethyl cellulose (Pb(CMC)(2)), which contains salicyl fluorones (THBF), Pb(CMC)(2)-THBF, were synthesized by sol-gel method. Pb(CMC)(2)-THBF can emit intense and stable solid substrate room temperature phosphorescence (SS-RTP) on filter paper. EDTA can chelate the Pb(2+) in Pb(CMC)(2)-THBF, causing it decompose into aqueous soluble components PbY(2-), CMC(-) and THBF, and these components can react with Hg(2+) to form (CMC)(2)Hg-THBF, causing decrease of phosphorescence intensity. Based on the facts above, a new method for the determination of trace mercury by SS-RTP quenching method was established. The linear range of this method is 2.0-40.0fgspot(-1) (5.0-100.0pgml(-1)) of Hg(2+), with a detection limit (LD) of 0.26fgspot(-1), and the regression equation of working curve is [Formula: see text] (fgspot(-1), 0.4mul spot(-1)), r = 0.9994. This method has been applied to the determination of trace mercury in water sample with satisfactory results. The mechanism of SS-RTP emission is also discussed.