An electrochemically switched smart surface for peptide immobilization and conformation control.

We report an electrochemically switched smart surface for controlled peptide immobilization and conformation control. This dynamic surface is based on self-assembled monolayers (SAMs) containing surface-bound trimethoxybenzene moieties, which can undergo electrochemically modulated surface activation to be stepwisely converted to two catechol derivatives. This new smart surface can be used to realize stepwise immobilization of a peptide, and more importantly, to control peptide conformation on a surface. We demonstrate herein that with one electrochemical activation step, a linear peptide containing an RGD sequence can be attached onto the SAMs. With the subsequence activation step, the attached linear RGD peptide can be converted into cyclic conformation. The SAMs bounded with linear and cyclic RGD exhibit different adhesion behaviors to fibroblasts cells. The reaction procedure can be well-monitored by cyclic voltammetry (CV), electrochemical surface enhanced Raman microscopy (EC-SERS), and X-ray photoelectron spectroscopy (XPS). It is believed this robust smart surface can find wide applications in surface immobilization of bioactive moieties.

A highly selective fluorescence "turn-on" probe for Cu(II) based on reaction and its imaging in living cells.

A new oxidative C-O bond cleavage reaction-based probe FluHMPP was designed and prepared. FluHMPP displays excellent selective turn-on fluorescence response for Cu(II) in aqueous solution under visible light excitation. The cleavage products are fully characterized. Fluorescein fragment is further oxidized to highly fluorescent MFME (3'-O-methylfluorescein methyl ester), and benzyl ether of imine fragment has been transformed to carboxyl. Confocal microscopy experiments have demonstrated that FluHMPP could also be used in live cells for the detection of Cu(II).

Potentilla anserina polysaccharide alleviates cadmium-induced oxidative stress and apoptosis of H9c2 cells by regulating the MG53-mediated RISK pathway.

Oxidative stress plays a crucial role in cadmium (Cd)-induced myocardial injury. Mitsugumin 53 (MG53) and its mediated reperfusion injury salvage kinase (RISK) pathway have been demonstrated to be closely related to myocardial oxidative damage. Potentilla anserina L. polysaccharide (PAP) is a polysaccharide with antioxidant capacity, which exerts protective effect on Cd-induced damage. However, it remains unknown whether PAP can prevent and treat Cd-induced cardiomyocyte damages. The present study was desgined to explore the effect of PAP on Cd-induced damage in H9c2 cells based on MG53 and the mediated RISK pathway. For in vitro evaluation, cell viability and apoptosis rate were analyzed by CCK-8 assay and flow cytometry, respectively. Furthermore, oxidative stress was assessed by 2',7'-dichlorodihydrofluorescein diacetate (DCFH-DA) staining and using superoxide dismutase (SOD), catalase (CAT), and glutathione/oxidized glutathione (GSH/GSSG) kits. The mitochondrial function was measured by JC-10 staining and ATP detection assay. Western blot was performed to detect the expression of proteins related to MG53, the RISK pathway, and apoptosis. The results indicated that Cd increased the levels of reactive oxygen species (ROS) in H9c2 cells. Cd decreased the activities of SOD and CAT and the ratio of GSH/GSSG, resulting in decreases in cell viability and increases in apoptosis. Interestingly, PAP reversed Cd-induced oxidative stress and cell apoptosis. Meanwhile, Cd reduced the expression of MG53 in H9c2 cells and inhibited the RISK pathway, which was mediated by decreasing the ratio of p-AktSer473/Akt, p-GSK3ßSer9/GSK3ß and p-ERK1/2/ERK1/2. In addition, Cd impaired mitochondrial function, which involved a reduction in ATP content and mitochondrial membrane potential (MMP), and an increase in the ratio of Bax/Bcl-2, cytoplasmic cytochrome c/mitochondrial cytochrome c, and Cleaved-Caspase 3/Pro-Caspase 3. Importantly, PAP alleviated Cd-induced MG53 reduction, activated the RISK pathway, and reduced mitochondrial damage. Interestingly, knockdown of MG53 or inhibition of the RISK pathway attenuated the protective effect of PAP in Cd-induced H9c2 cells. In sum, PAP reduces Cd-induced damage in H9c2 cells, which is mediated by increasing MG53 expression and activating the RISK pathway.

A retrievable and highly selective fluorescent probe for monitoring sulfide and imaging in living cells.

A novel selective fluorescent chemosensor based on an 8-hydroxyquinoline-appended fluorescein derivative (L1) was synthesized and characterized. Once combined with Cu(2+), it displayed high specificity for sulfide anion. Among the various anions, only sulfide anion induced the revival of fluoresecence of L1, which was quenched by Cu(2+), resulting in "off-on"-type sensing of sulfide anion. What's more, the sensor was retrievable to indicate sulfide anions with Cu(2+), and S(2-), in turn, increased. With the addition of Cu(2+), compound L1 could give rise to a visible pink-to-yellow color change and green fluorescence quenching. The resulting yellow solution could change to pink and regenerate to green fluorescence immediately upon the addition of sulfide anion; however, no changes were observed in the presence of other anions, including CN(-), P(2)O(7)(4-), and other forms of sulfate, making compound L1 an extremely selective and efficient sulfide chemosensor. The signal transduction occurs via reversible formation-separation of complex L1Cu and CuS. What's more, the biological imaging study has demonstrated that the chemosensor can detect sulfur anions in biological systems at a relatively low concentration.

Selective off-on fluorescent chemosensor for detection of Fe3+ ions in aqueous media.

A Fe(3+) chemosensor L1 was successfully synthesized with a quinoline moiety bound to rhodamine 6G hydrazide. The sensor L1 shows high selectivity and sensitivity to Fe(3+) in aqueous solution in the presence of other trace metal ions in organisms, abundant cellular cations and prevalent toxic metal ions in the environment. In addition, biological imaging and micro computed tomography (MCT) technology studies have demonstrated that L1 could act as a turn-on fluorescent chemosensor for Fe(3+) in living cells.

Potentilla anserine L. polysaccharide protects against cadmium-induced neurotoxicity.

Cadmium is a toxic metal that can damage the brain and other organs. This study aimed to explore the protective effects of Potentilla anserine L. polysaccharide (PAP) against CdCl2-induced neurotoxicity in N2a and SH-SY5Y cells and in the cerebral cortex of BALB/c mice. In addition, we aimed to identify the potential mechanisms underlying these protective effects. Relative to CdCl2 treatment alone, pretreatment with PAP prevented the reduction in cell viability evoked by CdCl2, decreased rates of apoptosis, promoted calcium homeostasis, decreased ROS accumulation, increased mitochondrial membrane potential, inhibited cytochrome C and AIF release, and prevented the cleavage of caspase-3 and PARP. In addition, PAP significantly decreased the CdCl2-induced phosphorylation of CaMKII, Akt, and mTOR. In conclusion, PAP represents a potential therapeutic agent for the treatment of Cd-induced neurotoxicity, functioning in part via attenuating the activation of the mitochondrial apoptosis pathway and the Ca2+-CaMKII-dependent Akt/mTOR pathway.

Biocompatible hydroxyapatite nanoparticles as a redox luminescence switch.

A redox luminescence switch was prepared by doping hydroxyapatite nanoparticles with CePO(4):Tb. The resulting multifunctional material exhibits good biocompatibility, biological affinity, and potential drug-carrying capability. The luminescent hydroxyapatite nanoparticles may find important applications in biomedical diagnostics, drug delivery, and biological sensors.

Potentilla anserine L. polysaccharide inhibits cadmium-induced neurotoxicity by attenuating autophagy.

Cadmium (Cd), a heavy metal with cytotoxicity, can activate autophagy. This study aimed to explore the effects and mechanisms of Potentilla anserine L. polysaccharide (PAP) on autophagy in N2a cells, primary neurons, and the brain of BALB/c mice exposed to Cd. The CCK-8 assay results showed that the cell viability decreased and the number of acidic vesicular organelles, autophagic vacuoles, lysosomes, and dysfunctional mitochondria increased in the cytoplasm of Cd-exposed N2a cells and primary neurons, as revealed by acridine orange staining, monodansylcadaverine staining, and transmission electron microscopy. PAP mitigated Cd-induced neuronal death and characteristic changes in autophagy. The expression of LC3 IILC3 II, Bcl-2, p62, Beclin-1, and PI3K class III was examined by Western blot analysis. Furthermore, the PI3K inhibitor (LY294002 or 3-MA) and/or PAP reversed the Cd-induced upregulated expression of LC3 II, Beclin-1, and PI3K class III, with a synergy between PI3K inhibitor and PAP against Cd-induced autophagy. The findings suggested that PAP partially prevented Cd-induced autophagic cell death in neurons by inhibiting the PI3K class III/Beclin-1 signaling pathway in vitro and in vivo.

Apoptotic effect of organophosphorus insecticide chlorpyrifos on mouse retina in vivo via oxidative stress and protection of combination of vitamins C and E.

Organophosphorus insecticide poisoning is widely investigated, and a growing number of evidence indicates its effects to cause ocular lesions, but the mechanisms of its ocular effects are not well elucidated. Here, effects of organophosphorus insecticide chlorpyrifos on mouse retina in vivo and protection of combination of vitamins C and E were reported. Cell apoptosis, lipid peroxidation and DNA damage were increased, and activities of antioxidant enzymes superoxide dismutase, catalase and glutathione peroxidase were decreased in retina of chlorpyrifos-administrated mice (63mg/kg, single treatment, via oral gavage). Pretreatment of combination of antioxidants vitamin C (250mg/kg) and vitamin E (150mg/kg) (once daily for 6 days, hypodermic injecting) significantly attenuated these effects of chlorpyrifos, demonstrating oxidative stress was involved in chlorpyrifos-induced cell apoptosis in mouse retina. Moreover, chlorpyrifos treatment inhibited acetylcholinesterase activity and promoted [Ca(2+)](i) level in mouse retinal cells, which were also attenuated by combination of vitamins C and E. These results may have implications for treatment of organophosphorus insecticide poisoning in retina with combination of vitamins C and E.

[Erythrocyte immunologic function effects of Patrinia scabra extracts by macroporous adsorptive resins on mice burdened transplanted tumor].

OBJECTIVE: To research the erythrocyte immunoregulation effects of Patrinia scabra extracts by macroporous adsorptive resins on mice burdened transplanted tumor. METHODS: Extracts of Patrinia scabra Bunge were separated by macroporous adsorptive resins, ingredients were analysised. Mice burdened transplanted tumor were given extracted drugs. Life prolongation rate was observed, erythrocyte immunologic function and the CD35, CD44s contents of red blood cell were evaluated. RESULTS: Polysaccharide and saponin accounted for 8.4% and 48.4%. Extracts could porolong life expectancy of mice, improve erythrocyte immunolgic function and increase the CD35 and CD44s contents of red blood cell. CONCLUSION: Extracts of Patrinia scabra Bunge by macroporous adsorptive resins have erythrocyte immunoregulation effects on mice burdened transplanted tumor.

Protective effect of Potentilla anserina polysaccharide on cadmium-induced nephrotoxicity in vitro and in vivo.

The aim of this research was to investigate the antioxidant and anti-apoptotic activities of Potentilla anserina polysaccharide (PAP) on kidney damage induced by cadmium (Cd) in vitro and in vivo. PAP has been suggested to have anti-oxidation, anti-apoptosis, immunoregulation, antimicrobial, antitussive, and expectorant abilities. In this study, PAP was extracted and the major components of PAP were analyzed. It was shown that PAP pretreatment remarkably improved redox homeostasis, both in human embryonic kidney 293 (HEK293) cells and in BALB/c mice. Administration of PAP attenuated the mitochondrial dysfunction, degeneration, and fibrosis of kidney induced by Cd. Furthermore, PAP exhibited anti-apoptotic activity, which involved regulating both the mitochondria-mediated intrinsic apoptotic pathway and the death receptor-initiated extrinsic pathway. These results suggest that PAP is a potential therapeutic agent for Cd-induced nephrotoxicity.

[Effects of fuzheng yiliu granules on apoptotic rate and mitochondrial membrane potential of hepatocellular carcinoma cell line H22 from mice].

OBJECTIVE: To evaluate the effects of Fuzheng Yiliu Granules (FZYLG) on apoptotic rate and mitochondrial membrane potential (Delta psi m) of hepatocellular carcinoma cell line H22 from mice. METHODS: Forty-eight mice inoculated with H22 cells were randomly divided into four groups: untreated group, cyclophosphamide-treated group, high-dose FZYLG-treated group and low-dose FZYLG-treated group. After 14 days of corresponding treatment, H22 cells in each group were stained with propidium iodide, and the apoptotic rates were detected by flow cytometry (FCM). The rhodamine 123 was used as a fluorescence probe to label the H22 cells, and the fluorescence intensities were observed with laser scanning confocal microscope. The fluorescence intensity of H22 cells indicated the Delta psi m of H22 cells. RESULTS: FZYLG could significantly increase the apoptotic rate while reduce the Delta psi m of H22 cells from mice as compared with those in the untreated group. CONCLUSION: The antitumor effects of FZYLR on H22 cells from mice are related to decreasing the Delta psi m and then inducing the apoptosis of the H22 cells.

Proanthocyanidin from grape seeds enhances anti-tumor effect of doxorubicin both in vitro and in vivo.

The purpose of this study was to investigate the synergistic anti-tumor effect of proanthocyanidin (PA) and doxorubicin (DOX) on K562, A549 and CNE cells in vitro and experimental transplantation Sarcoma 180 (S180) and Hepatoma 22 (H22) in vivo and to explore the mechanism of its action. PA 12.5 approximately 100 mg/l inhibited proliferation of K562, A549, and CNE cells in vitro in a time- and concentration-dependent manner as determined by the microculture tetrazolium (MTT) assay. A combination of PA 12.5, 25 mg/l with DOX 0.01 approximately 1 mg/l treatment synergistically inhibited proliferation of K562, A549, and CNE cells with decreased IC50 values. Under the confocal laser scanning microscope, intracellular DOX, Ca2+, and Mg2+ concentrations were greatly increased whereas pH value and mitochondrial membrane potential were markedly reduced in K562 cells after treatment with a combination of PA plus DOX. At the same time, K562 cells showed morphological changes of apoptosis following treatment with PA plus DOX, and the administration of PA 25 mg/l plus DOX 0.3 mg/l for 24 h resulted in a significant increase in the percentage of apoptosis by flow cytometry as compared with DOX 0.3 mg/l alone (p < 0.05). In vivo experiments showed that a combination of PA 200 mg/kg i.g. with DOX 2 mg/kg i.p. treatment displayed an inhibitory effect on the growth of transplantation tumor S180 and H22 in mice compared with the DOX only group (p < 0.01). Taken together, these results suggest that PA enhances the DOX-induced anti-tumor effect and its mechanism is attributed to the promotion of DOX-induced apoptosis through increasing intracellular DOX, Ca2+ and Mg2+ concentrations, and reducing pH value and mitochondrial membrane potential.

A coumarin-based two-photon probe for hydrogen peroxide.

A new fluorescence probe was developed for hydrogen peroxide (H2O2) detection based on donor-excited photo induced electron transfer (D-PET) mechanism, together with the benzil as a quenching and recognizing moiety. The benzil could convert to benzoic anhydride via a Baeyer-Villiger type reaction in the presence of H2O2, followed by hydrolysis of benzoicanhydride to give benzoic acid, and the fluorophore released. The probe was synthesized by a 6-step procedure starting from 4-(diethylamino)salicylaldehyde. A density functional theory (DFT) calculation was performed to demonstrate that the benzil was a fluorescence quencher. The probe was evaluated in both one-photon and two-photon mode, and it exhibited high selectivity toward H2O2 over other reactive oxygen species and high sensitivity with a detection limit of 0.09 µM. Furthermore, the probe was successfully applied to cell imaging of intracellular H2O2 levels with one-photon microscopy and two-photon microscopy. The superior properties of the probe made it of great potential use in more chemical and biological researches.

Rational design of small indolic squaraine dyes with large two-photon absorption cross section.

Small organic dyes with large two-photon absorption (TPA) cross sections (δ) are more desirable in many applications compared with large molecules. Herein, we proposed a facile theoretical method for the fast screening of small organic molecules as potential TPA dyes. This method is based on a theoretical analysis to the natural transition orbitals (NTOs) directly associated with the TPA transition. Experimental results on the small indolic squaraine dyes (ISD) confirmed that their TPA cross sections is strongly correlated to the delocalization degree of the NTOs of the S2 excited states. Aided by this simple and intuitive method, we have successfully designed and synthesized a small indolic squaraine dye (ISD) with a remarkable δ value above 8000 GM at 780 nm. The ISD dye also exhibits a high singlet oxygen generation quantum yield about 0.90. The rationally designed TPA dye was successfully applied in both two-photon excited fluorescence cell imaging and in vivo cerebrovascular blood fluid tracing.

Arsenic trioxide inhibits p-glycoprotein expression in multidrug-resistant human leukemia cells that overexpress the MDR1 gene.

OBJECTIVE: To investigate the effects of arsenic trioxide (As(2)O(3)) on the apoptosis and p-glycoprotein (P-gp) expression of multidrug-resistant human leukemia cells. METHODS: Human multidrug-resistant leukemia cell line K562/ADM overexpressing the MDR1 gene, was used as the target cells. The cell proliferating activity was assessed using the MTT colorimetric assay. Cytomorphology was investigated under light, confocal and electron microscopes. DNA fragmentation was examined using agarose gel electrophoresis, while p-gp expression, cell cycle status and sub-G1 cells were determined using flow cytometry. RESULTS: Zero point five to 20 micromol/L As(2)O(3) inhibited the proliferation of K562/ADM cells, and K562/ADM cells were more sensitive to As(2)O(3) than the parental K562 cells. As(2)O(3)-induced apoptosis of K562/ADM cells was determined by the observance of typical morphological changes and the appearance of DNA ladder and sub-G1 cell populations. As(2)O(3) significantly inhibited the P-gp expression of K562/ADM cells, and synergistically enhanced the sensitivity of the drug-resistant cells to adriamycin. CONCLUSIONS: As(2)O(3) induces growth-inhibition and apoptosis, down-regulates P-gp expression and exerts a synergistic effect in combination with adriamycin in multidrug-resistant leukemia cells.

Gelatin functionalized graphene oxide for mineralization of hydroxyapatite: biomimetic and in vitro evaluation.

We report a facile modification of graphene oxide (GO) by gelatin to mimic charged proteins present in the extracellular matrix during bone formation. The bioinspired surface of GO-gelatin (GO-Gel) composite was used for biomimetic mineralization of hydroxyapatite (HA). A detailed structural and morphological characterization of the mineralized composite was performed. Additionally, MC3T3-E1 cells were cultured on the GO-Gel surfaces to observe various cellular activities and HA mineralization. Higher cellular activities such as cell adhesion, cell proliferation, and alkaline phosphatase activity (ALP) were observed on the GO-Gel surface compared with the GO or glass surface. The increase of ALP confirms that the proposed GO-Gel promotes the osteogenic differentiation of MC3T3-E1 cells. Moreover, the evidence of mineralization evaluated by scanning electron microscopy (SEM) and alizarin red staining (ARS) corroborate the idea that a native osteoid matrix is ultimately deposited. All these data suggest that the GO-Gel hybrids will have great potential as osteogenesis promoting scaffolds for successful application in bone surgery.

Biomimetic and cell-mediated mineralization of hydroxyapatite by carrageenan functionalized graphene oxide.

In bone tissue engineering, it is imperative to design multifunctional biomaterials that can induce and assemble bonelike apatite that is close to natural bone. In this study, graphene oxide (GO) was functionalized by carrageenan. The resulting GO-carrageenan (GO-Car) composite was further used as a substrate for biomimetic and cell-mediated mineralization of hydroxyapatite (HA). It was confirmed that carrageenan on the GO surface facilitated the nucleation of HA. The observation of the effect of the GO-Car on the adhesion, morphology, and proliferation of MC3T3-E1 cells was investigated. In vitro studies clearly show the effectiveness of GO-Car in promoting HA mineralization and cell differentiation. The results of this study suggested that the GO-Car hybrid will be a promising material for bone regeneration and implantation.

A coumarin-derived fluorescent chemosensor for selectively detecting Cu²âº: synthesis, DFT calculations and cell imaging applications.

A novel coumarin-based fluorescent probe L ((4E)-4-((7-hydroxy-4-methyl-2-oxo-2H-chromen-8-yl) methyleneamino)-1,2-dihyydro-2,3-dimethyl-1-phenylpyrazol-5-one) has been developed as a simple and efficient chemosensor which exhibits a significant fluorescence reduction in the presence of metal cations. This sensor exhibits high selectivity and sensitivity toward Cu(2+) over other common cations. The mechanism for detecting copper was evaluated by time-dependent density functional theory (TD-DFT) calculations and the coordination mode was also confirmed by density functional theory (DFT) calculations. Furthermore, results of cell imaging in this study indicate that this new probe may be useful for detection and monitoring of Cu(2+) in biological applications.

[Effects of non-saccharomyces albicans metabolic products on the proliferation of human umbilical vein endothelial cell ECV304].

OBJECTIVE: To evaluate the effects of non-Saccharomyces albicans metabolic products on the cell cycle distribution and proliferation of human umbilical vein endothelial cell ECV304 cells in vitro. METHODS: The parallel dilution supernatant of Saccharomyces tropicalis, Saccharomyces krusei and Saccharomyces glabrata were prepared, and 1, 4, 16-fold(s) diluted concentration and control group were set up. The line of human umbilical vein endothelial cell ECV304 was cultured in vitro and treated by non-Saccharomyces albicans supernatant. The proliferous effect of ECV304 induced by non-Saccharomyces albicans supernatant after 24, 48, 72 h was detected by the methods of MTT, and the changes of cell density and cycle after 48 h were investigated by inverted microscope and flow cytometry. RESULTS: At the 24th hour, all of the higher concentration (1-fold) of non-Saccharomyces albicans supernatant and the 4-folds diluted Saccharomyces krusei could promote ECV304 proliferation(P < 0.05). After adding various non-Saccharomyces albicans supernatant at 48h and 72th hour, Saccharomyces krusei supernatant and Saccharomyces glabrata supernatant significantly increased proliferation rate of ECV304, while Saccharomyces tropicalis supernatant group showed no significant change no matter which concentration was tested. At 48th hour after adding the non-Saccharomyces albicans supernatant, the ECV304 cells density treated by Saccharomyces krusei supernatant and Saccharomyces glabrata supernatant were significantly higher under the inverted microscope. The G0/G1 population of ECV304 cells decreased while cell proliferation index (PI) increased after incubated with Saccharomyces krusei supernatant and Saccharomyces glabrata supernatant for 48 hours (P < 0.05). Saccharomyces tropicalis group showed no significant change (P > 0.05). CONCLUSION: The metabolic products of Sacharoymces krusei and Saccharomyces glabrata could induce proliferation of ECV304 cell, which suggests non-Saccharomyces albicans should be undergone more attention clinically in detection and treatment.