Palmdelphin Deficiency Evokes NF-κB Signaling in Valvular Endothelial Cells and Aggravates Aortic Valvular Remodeling.

Palmd-deficient mice of advanced age manifest increased aortic valve peak velocity, thickened aortic valve leaflets, and excessive extracellular matrix deposition, which are key features of calcific aortic valve disease. PALMD is predominantly expressed in endothelial cells of aortic valves, and PALMD-silenced valvular endothelial cells are prone to oscillatory shear stress-induced endothelial-to-mesenchymal transition. Mechanistically, PALMD is associated with TNFAIP3 interaction protein 1, a binding protein of TNFAIP3 and IKBKG in NF-κB signaling. Loss of PALMD impairs TNFAIP3-dependent deubiquitinating activity and promotes the ubiquitination of IKBKG and subsequent NF-κB activation. Adeno-associated virus-mediated PALMD overexpression ameliorates aortic valvular remodeling in mice with calcific aortic valve disease, indicating protection.

IL-17A induces valvular endothelial inflammation and aggravates calcific aortic valve disease.

Calcific aortic valve disease (CAVD) is an aging related disease characterized by inflammation and fibrocalcific remodeling. IL-17A is a key cytokine associated with pathophysiology of inflammatory and fibrotic disease. Previous studies showed accumulation of IL-17A-producing T helper lymphocytes in human calcified aortic valves and significantly elevated IL-17RA expression in calcified valves. However, the role of IL-17A signaling in the initiation and development of CAVD is still unclear. In this study, by analyzing public transcriptome databases, we found that IL-17A-IL-17RA signaling is activated in calcified valves. Gene expression analysis revealed significantly increased IL-17A, IL-17RA, and RUNX2 expression in calcified human aortic valves compared to in non-calcified valves, and the expression of IL-17A and IL-17RA were positively correlated with RUNX2 expression. A 5/6 nephrectomy was performed in Apoe-/- (Apoe knockout) mice to establish a CAVD mouse model. IL-17A-neutralizing antibodies significantly reduced valve calcium deposition and decreased expression of RUNX2 in aortic valves. Immunofluorescence staining of human aortic valves and qRT-PCR analysis of primary aortic valve cells revealed abundant expression of IL-17RA in valvular endothelial cells (VECs). RNA sequencing indicated that IL-17A promoted the activation of inflammatory signaling pathways in VECs. Furthermore, qRT-PCR and cytometric bead array analysis confirmed that IL-17A promoted the expression or secretion of inflammatory cytokines IL-6 and IL-1ß, chemokines CXCL2 and CXCL8, and fibrosis-related gene COL16A1. Our findings indicate that elevated IL-17A in CAVD may promote valve inflammation, fibrosis, and calcification by inducing endothelial activation and inflammation. Targeting IL-17A-IL-17RA signaling may be a potential therapeutic strategy for CAVD.

Biodegradation of arsenobetaine to inorganic arsenic regulated by specific microorganisms and metabolites in mice.

Arsenobetaine (AsB) is a primary arsenic (As) compound found in marine organisms. However, in mammals, the metabolic mechanism of AsB remains indistinct. Therefore, in this study, we investigated the biotransformation and regulatory mechanism of AsB, particularly the biodegradation process, in a mouse model to assess the underlying health hazards of AsB. We studied the biotransformation process of AsB in mice through the food chain [AsB feed-marine fish (Epinephelus fuscoguttatus)-mice (Mus musculus)]. Our results showed the significant bioaccumulation of total As, AsB, and, in particular, arsenate [As(V)] through biodegradation in mice tissues. As the abundance of Staphylococcus and Blautia (phylum, Firmicutes) increased, the expression of aqp7 (absorption) and methyltransferase (as3mt) (methylation) was upregulated. In contrast, the expression of S-adenosyl methionine (sam) (methylation) was downregulated. These findings suggest that demethylation and methylation occurred simultaneously in the intestines, with demethylation capacity being greater than that of methylation. Furthermore, Firmicutes such as Staphylococcus and Blautia showed a significant inverse relationship with arachidonic acid, choline, and sphingosine. Gene, microbiome, and metabolomics analyses indicated that Staphylococcus and Blautia and arachidonic acid, choline, and sphingosine participated in the degradation of AsB to As(V) in mouse intestines. Therefore, long-term AsB ingestion through marine fish consumption could cause potential health hazards in humans.

Magnesium dicarboxylates promote the prenylation of phenolics that is extended to the total synthesis of icaritin.

The prenylation of phenolic substrates promoted by magnesium dicarboxylates was developed. An investigation of the scope demonstrated that substrates with electron-donating group(s) gave better yields than those with electron-withdrawing group(s). Although the conversions of all substrates were higher in MeCN than in DMF, DMF was still the favorable solvent for polyphenolic substrates since MeCN would cause the generation of cyclized by-products (6) and reduce the yield of 3. The regio-selectivity of ortho- vs. para-prenylation (3'vs.3'') for those para-unoccupied substrates was also solvent dependant. DMF produced mainly ortho-products but with poor conversions. On the other hand, MeCN generated mainly para-products, along with minor ortho-products. Mechanistic study of the prenylation provided evidence for the nucleophilic addition/substitution of the phenolic substrate to the alkyl halide in the presence of the magnesium dicarboxylates. The proto application of this method in the total synthesis of icaritin through the prenylation of 2,4,6-trihydroxyacetophenone, followed by the reaction with benzaldehyde to afford the flavonol, was successful, with a total yield of 33%.

Identification of Pyruvate Carboxylase as the Cellular Target of Natural Bibenzyls with Potent Anticancer Activity against Hepatocellular Carcinoma via Metabolic Reprogramming.

Cancer cell proliferation in some organs often depends on conversion of pyruvate to oxaloacetate via pyruvate carboxylase (PC) for replenishing the tricarboxylic acid cycle to support biomass production. In this study, PC was identified as the cellular target of erianin using the photoaffinity labeling-click chemistry-based probe strategy. Erianin potently inhibited the enzymatic activity of PC, which mediated the anticancer effect of erianin in human hepatocellular carcinoma (HCC). Erianin modulated cancer-related gene expression and induced changes in metabolic intermediates. Moreover, erianin promotes mitochondrial oxidative stress and inhibits glycolysis, leading to insufficient energy required for cell proliferation. Analysis of 14 natural analogs of erianin showed that some compounds exhibited potent inhibitory effects on PC. These results suggest that PC is a cellular target of erianin and reveal the unrecognized function of PC in HCC tumorigenesis; erianin along with its analogs warrants further development as a novel therapeutic strategy for the treatment of HCC.

[Arachidonic acid Alox15/12-HETE signaling inhibits vascular calcification].

This study aims to explore the effects of arachidonic acid lipoxygenase metabolism in vascular calcification. We used 5/6 nephrectomy and high-phosphorus feeding to establish a model of vascular calcification in mice. Six weeks after nephrectomy surgery, vascular calcium content was measured, and Alizarin Red S and Von Kossa staining were applied to detect calcium deposition in aortic arch. Control aortas and calcified aortas were collected for mass spectrometry detection of arachidonic acid metabolites, and active molecules in lipoxygenase pathway were analyzed. Real-time quantitative PCR was used to detect changes in the expression of lipoxygenase in calcified aortas. Lipoxygenase inhibitor was used to clarify the effect of lipoxygenase metabolic pathways on vascular calcification. The results showed that 6 weeks after nephrectomy surgery, the aortic calcium content of the surgery group was significantly higher than that of the sham group (P < 0.05). Alizarin Red S staining and Von Kossa staining showed obvious calcium deposition in aortic arch from surgery group, indicating formation of vascular calcification. Nine arachidonic acid lipoxygenase metabolites were quantitated using liquid chromatography/mass spectrometry (LC-MS) analysis. The content of multiple metabolites (12-HETE, 11-HETE, 15-HETE, etc.) was significantly increased in calcified aortas, and the most abundant and up-regulated metabolite was 12-HETE. Furthermore, we examined the mRNA levels of metabolic enzymes that produce 12-HETE in calcified blood vessels and found the expression of arachidonate lipoxygenase-15 (Alox15) was increased. Blocking Alox15/12-HETE by Alox15 specific inhibitor PD146176 significantly decreased the plasma 12-HETE content, promoted calcium deposition in aortic arch and increased vascular calcium content. These results suggest that the metabolism of arachidonic acid lipoxygenase is activated in calcified aorta, and the Alox15/12-HETE signaling pathway may play a protective role in vascular calcification.

MicroRNA-223-3p inhibits vascular calcification and the osteogenic switch of vascular smooth muscle cells.

Vascular calcification is the ectopic deposition of calcium hydroxyapatite minerals in arterial wall, which involves the transdifferentiation of vascular smooth muscle cells (VSMCs) toward an osteogenic phenotype. However, the underlying molecular mechanisms regulating the VSMC osteogenic switch remain incompletely understood. In this study, we examined the roles of microRNAs (miRNAs) in vascular calcification. miRNA-seq transcriptome analysis identified miR-223-3p as a candidate miRNA in calcified mouse aortas. MiR-223-3p knockout aggravated calcification in both medial and atherosclerotic vascular calcification models. Further, RNA-seq transcriptome analysis verified JAK-STAT and PPAR signaling pathways were upregulated in both medial and atherosclerotic calcified aortas. Overlapping genes in these signaling pathways with predicted target genes of miR-223-3p derived from miRNA databases, we identified signal transducer and activator of transcription 3 (STAT3) as a potential target gene of miR-223-3p in vascular calcification. In vitro experiments showed that miR-223-3p blocked interleukin-6 (IL-6)/STAT3 signaling, thereby preventing the osteogenic switch and calcification of VSMCs. In contrast, overexpression of STAT3 diminished the effect of miR-223-3p. Taken together, the results indicate a protective role of miR-223-3p that inhibits both medial and atherosclerotic vascular calcification by regulating IL-6/STAT3 signaling-mediated VSMC transdifferentiation.

Metabolic Activation and Covalent Protein Binding of Berberrubine: Insight into the Underlying Mechanism Related to Its Hepatotoxicity.

INTRODUCTION: Berberrubine (BRB), an isoquinoline alkaloid, is a major constituent of medicinal plants Coptis chinensis Franch or Phellodendron chinense Schneid. BRB exhibits various pharmacological activities, whereas exposure to BRB may cause toxicity in experimental animals. METHODS: In this study, we thoroughly investigated the liver injury induced by BRB in mice and rats. To explore the underlying mechanism, a study of the metabolic activation of BRB was conducted. Furthermore, covalent modifications of cysteine residues of proteins were observed in liver homogenate samples of animals after exposure to BRB, by application of an exhaustive proteolytic digestion method. RESULTS: It was demonstrated that BRB-induced hepatotoxicities in a time- and dose-dependent manner, based on the biochemical parameters ALT and AST. H&E stained histopathological examination showed the occurrence of obvious edema in liver of mice after intraperitoneal (i.p.) administration of BRB at a single dose of 100 mg/kg. Slight hepatotoxicity was also observed in rats given the same doses of BRB after six weeks of gavage. As a result, four GSH adducts derived from reactive metabolites of BRB were detected in microsomal incubations with BRB fortified with GSH as a trapping agent. Moreover, four cys-based adducts derived from reaction of electrophilic metabolites of BBR with proteins were found in livers. CONCLUSION: These results suggested that the formation of protein adducts originating from metabolic activation of BRB could be a crucial factor of the mechanism of BRB-induced toxicities.

Mechanisms of potentially toxic metal removal from biogas residues via vermicomposting revealed by synchrotron radiation-based spectromicroscopies.

Biogas residues (BR) contaminated with potentially toxic metals pose environmental risks to soils and food chains, and strategies are needed to decrease the concentration and bioavailability of potentially toxic metals in BR. Here, metal fractions and removal mechanisms were quantified by synchrotron radiation-based Fourier transform infrared and micro X-ray fluorescence spectromicroscopies on BR and earthworms subject to vermicomposting. Vermicomposting resulted in decreases in concentrations of potentially toxic metals in BR and increases in metal removal efficiencies due to uptake by earthworms. Prior to vermicomposting, Zn, Cu and Pb were associated with N-H, O-H, aromatic C, aliphatic C, and amide functional groups, but following maturation during vermicomposting, metals were associated with N-H, O-H, aliphatic C and polysaccharide functional groups. Following vermicomposting, Zn and Cu were mainly distributed in the dermal portions of earthworms, whereas Pb was more homogeneously distributed among the inner and outer portions of the earthworms, revealing that different metals may have different uptake routes. These findings provide a new strategy for safe utilization of BR by using earthworms via vermicomposting to remove potentially toxic metals and in situ insights into how metals binding and distribution characteristics in BR and earthworms during compost and vermicomposting processes.

Ultrasensitive Electrochemical DNA Biosensor Based on a Label-Free Assembling Strategy Using a Triblock polyA DNA Probe.

Multiblock DNA probe attracted a large amount of scientific attention, for the development of multitarget biosensor and improved specificity/sensitivity. However, the development of multiblock DNA probes highly relied on the chemical synthesis of organic linkers or nanomaterials, which limited their practicability and biological compatibility. In this work, we developed a label-free assembling strategy using a triblock DNA capture probe, which connects two DNA probes with its intrinsic polyA fragment (probe-PolyA-probe, PAP). The middle polyA segment has a high affinity to the gold electrode surface, leading to excellent reproducibility, stability, and regeneration of our biosensor. Two flanking capture probes were tandemly co-assembled on the electrode surface with consistent spatial relationship and exactly the same amount. When combined with the target DNA, the hybridization stability was improved, because of the strong base stacking effect of two capture probes. The sensitivity of our biosensor was proved to be 10 fM, with a wide analysis range between 10 fM to 1 nM. Our PAP-based biosensor showed excellent specificity when facing mismatched DNA sequences. Even single nucleotide polymorphisms can be distinguished by each probe. The excellent practicability of our biosensor was demonstrated by analyzing genomic DNA both with and without PCR amplification.

A room-temperature interfacial approach towards iron/nitrogen co-doped fibrous porous carbons as electrocatalysts for the oxygen reduction reaction and Zn-Air batteries.

The development of nonprecious and efficient catalysts to boost the oxygen reduction reaction (ORR) is imperative. However, the majority of previously reported approaches suffered from a complicated fabrication procedure, both time consuming and difficult to scale up. Herein, large-scale iron ion embedded polyaniline fibers were successfully fabricated as precursors for preparing iron/nitrogen co-doped fibrous porous carbons (Fe/NPCFs) through an interfacial engineering strategy at room temperature. As ORR electrocatalysts in an alkaline medium (0.1 M KOH), Fe/NPCFs display a positive half-wave potential of 0.827 V (vs. RHE), and high limited current density (up to 5.76 mA cm-2), which are better than those of commercial Pt/C (E1/2 = 0.815 V, JL = 5.47 mA cm-2). Also, Fe/NPCFs exhibit a high ORR catalysis activity (E1/2 = 0.632 V, JL = 5.07 mA cm-2) in acidic medium (0.5 M H2SO4). When used as an air cathode in a primary Zn-air battery, high power density (158.5 mW cm-2) and specific capacity (717.8 mA h g-1) can be easily achieved, outperforming the commercial Pt/C.

Nanodiamond autophagy inhibitor allosterically improves the arsenical-based therapy of solid tumors.

Arsenic trioxide (ATO) is a successful chemotherapeutic drug for blood cancers via selective induction of apoptosis; however its efficacy in solid tumors is limited. Here we repurpose nanodiamonds (NDs) as a safe and potent autophagic inhibitor to allosterically improve the therapeutic efficacy of ATO-based treatment in solid tumors. We find that NDs and ATO are physically separate and functionally target different cellular pathways (autophagy vs. apoptosis); whereas their metabolic coupling in human liver carcinoma cells remarkably enhances programmed cell death. Combination therapy in liver tumor mice model results in ~91% carcinoma decrease as compared with ~28% without NDs. Treated mice show 100% survival rate in 150 days with greatly reduced advanced liver carcinoma-associated symptoms, and ~80% of post-therapy mice survive for over 20 weeks. Our work presents a novel strategy to harness the power of nanoparticles to broaden the scope of ATO-based therapy and more generally to fight solid tumors.

Identification of the predictive genes for the response of colorectal cancer patients to FOLFOX therapy.

BACKGROUND: Colorectal cancer is a malignant tumor with high death rate. Chemotherapy, radiotherapy and surgery are the three common treatments of colorectal cancer. For early colorectal cancer patients, postoperative adjuvant chemotherapy can reduce the risk of recurrence. For advanced colorectal cancer patients, palliative chemotherapy can significantly improve the life quality of patients and prolong survival. FOLFOX is one of the mainstream chemotherapies in colorectal cancer, however, its response rate is only about 50%. METHODS: To systematically investigate why some of the colorectal cancer patients have response to FOLFOX therapy while others do not, we searched all publicly available database and combined three gene expression datasets of colorectal cancer patients with FOLFOX therapy. With advanced minimal redundancy maximal relevance and incremental feature selection method, we identified the biomarker genes. RESULTS: A Support Vector Machine-based classifier was constructed to predict the response of colorectal cancer patients to FOLFOX therapy. Its accuracy, sensitivity and specificity were 0.854, 0.845 and 0.863, respectively. CONCLUSION: The biological analysis of representative biomarker genes suggested that apoptosis and inflammation signaling pathways were essential for the response of colorectal cancer patients to FOLFOX chemotherapy.

Hydrogen Sulfide-Activatable Second Near-Infrared Fluorescent Nanoassemblies for Targeted Photothermal Cancer Therapy.

Near-infrared (NIR)-II fluorescence agents hold great promise for deep-tissue photothermal therapy (PTT) of cancers, which nevertheless remains restricted by the inherent nonspecificity and toxicity of PTT. In response to this challenge, we herein develop a hydrogen sulfide (H2S)-activatable nanostructured photothermal agent (Nano-PT) for site-specific NIR-II fluorescence-guided PTT of colorectal cancer (CRC). Our in vivo studies reveal that this theranostic Nano-PT probe is specifically activated in H2S-rich CRC tissues, whereas it is nonfunctional in normal tissues. Activation of Nano-PT not only emits NIR-II fluorescence with deeper tissue penetration ability than conventional fluorescent probes but also generates high NIR absorption resulting in efficient photothermal conversion under NIR laser irradiation. Importantly, we establish NIR-II imaging-guided PTT of CRC by applying the Nano-PT agent in tumor-bearing mice, which results in complete tumor regression with minimal nonspecific damages. Our studies thus shed light on the development of cancer biomarker-activated PTT for precision medicine.

Black TiO2 based core-shell nanocomposites as doxorubicin carriers for thermal imaging guided synergistic therapy of breast cancer.

TiO2 nanomaterials have been widely used for anticancer drug carriers and UV/980 nm NIR triggered cancer synergistic platforms. However, traditional pure TiO2 nanocarriers encounter some serious drawbacks, such as low drug loading ability, limited tissue penetration of UV light, and heating effect of 980 nm NIR on normal tissue, which obstruct their further application in cancer treatment. To overcome those challenges, novel mesoporous silica (mSiO2) coated black TiO2 core-shell nanocomposites are designed and constructed as doxorubicin carriers for 808 nm NIR triggered thermal imaging guided photothermal therapy combined chemotherapy of breast cancer. Properties of the nanocomposites such as micro-morphology, size, drug loading ability and release, targeting performance, and therapy efficiency in vitro and in vivo were evaluated. The results indicated the core-shell nanocomposites with dramatically increased loading ability were pH-responsive/NIR-accelerated doxorubicin release nanocarriers and showed synergistic breast cancer treatment in vitro and in vivo. This study verifies that the newly prepared mSiO2 coated black TiO2 core-shell nanocarriers can overcome the limitations of traditional TiO2 nanocarriers and thus improve and broaden usage of TiO2 nanoparticles in nanomedicine.

[Clinical observation of superficial branch of radial artery wrist crease flap for repairing ring tissue defect of fingers].

Objective: To explore the effectiveness and operation method of the superficial branch of radial artery wrist crease flap for repair of ring tissue defect of the fingers. Methods: Between June 2013 and March 2016, the superficial branch of radial artery wrist crease flap was used to repair ring finger tissue defect in 20 cases (21 fingers). There were 14 males and 6 females with an average age of 39.3 years (range, 12-61 years). The causes included machine injury in 9 cases, traffic accident injury in 6 cases, heat inury in 2 cases, and avulsed injury in 3 cases. The index finger was involved in 6 cases, middle finger in 6 cases, ring finger in 3 cases, and little finger in 6 cases. Combined injuries included exposure of bone, tendon, vessel, and nerve. The mean time of injury to operation was 3 hours (range, 0.5-5.5 hours) in 17 patients undergoing emergency operation, and was 8.5 days (range, 7-10 days) in 3 patients undergoing selective operation. The superficial palmar branch of the radial artery from the flap was used for bridging proper digital artery. The donor site was directly sutured in 19 cases and was repaired by skin grafting in 1 case. Results: One case had blood blister at distal flap, which was cured after dressing change; the other flaps survived, and primary healing was obtained. Healing of incision at the donor site healed by first intention. The patients were followed up 6-24 months (mean, 12 months). The appearance, texture, and color of the flaps were satisfactory. The two-point discrimination ranged from 6 to 13 mm (mean, 9 mm) at 6 months after operation. According to the Chinese Medical Association Society of hand surgery of thumb and finger reconstruction function evaluation standard, the results were excellent in 13 cases, good in 4 cases, and fair in 3 cases; the excellent and good rate was 85%. Conclusion: The superficial branch of radial artery wrist crease flap is an ideal choice for the repair of ring tissue defect of the fingers.

808 nm-excited upconversion nanoprobes with low heating effect for targeted magnetic resonance imaging and high-efficacy photodynamic therapy in HER2-overexpressed breast cancer.

To avoid the overheating effect of excitation light and improve the efficacy of photodynamic therapy (PDT) of upconversion nanoplatform, a novel nanoprobe based on 808 nm-excited upconversion nanocomposites (T-UCNPs@Ce6@mSiO2) with low heating effect and deep penetration has been successfully constructed for targeted upconversion luminescence, magnetic resonance imaging (MRI) and high-efficacy PDT in HER2-overexpressed breast cancer. In this nanocomposite, photosensitizers (Ce6) were covalently conjugated inside of mesoporous silica to enhance the PDT efficacy by shortening the distance of fluorescence resonance energy transfer and to decrease the cytotoxicity by preventing the undesired leakage of Ce6. Compared with UCNPs@mSiO2@Ce6, UCNPs@Ce6@mSiO2 greatly promoted the singlet oxygen generation and amplified the PDT efficacy under the excitation of 808 nm laser. Importantly, the designed nanoprobe can greatly improve the uptake of HER2-positive cells and tumors by modifying the site-specific peptide, and the in vivo experiments showed excellent MRI and PDT via intravenous injection by modeling MDA-MB-435 tumor-bearing nude mice. Our strategy may provide an effective solution for overcoming the heating effect and improving the PDT efficacy of upconversion nanoprobes, and has potential application in visualized theranostics of HER2-overexpressed breast cancer.

Gd-based upconversion nanocarriers with yolk-shell structure for dual-modal imaging and enhanced chemotherapy to overcome multidrug resistance in breast cancer.

Multidrug resistance (MDR) of cancers is still a major challenge, and it is very important to develop visualized nanoprobes for the diagnosis and treatment of drug resistant cancers. In this work, we developed a multifunctional delivery system based on DOX-encapsulated NaYF4:Yb/Er@NaGdF4 yolk-shell nanostructures for simultaneous dual-modal imaging and enhanced chemotherapy in drug resistant breast cancer. Using the large pore volume of the nanostructure, the delivery system had a high loading efficiency and excellent stability. Also, an in vitro and in vivo toxicity study showed the good biocompatibility of the as-prepared yolk-shell nanomaterials. Moreover, by nanocarrier delivery, the uptake of DOX could be greatly increased in drug resistant MCF-7/ADR cells. Compared with free DOX, the as-prepared delivery system enhanced the chemotherapy efficacy against MCF-7/ADR cells, indicating the excellent capability for overcoming MDR. Furthermore, core-shell NaYF4:Yb/Er@NaGdF4 improved the upconversion luminescence (UCL) performance, and the designed delivery system could also be applied for simultaneous UCL and magnetic resonance (MR) imaging, which could be a good candidate as a dual-modal imaging nanoprobe. Therefore, we developed a multifunctional yolk-shell delivery system, which could have potential applications as a visualized theranostic nanoprobe to overcome MDR in breast cancer.

A heavy metal-associated protein (AcHMA1) from the halophyte, Atriplex canescens (Pursh) Nutt., confers tolerance to iron and other abiotic stresses when expressed in Saccharomyces cerevisiae.

Many heavy metals are essential for metabolic processes, but are toxic at elevated levels. Metal tolerance proteins provide resistance to this toxicity. In this study, we identified and characterized a heavy metal-associated protein, AcHMA1, from the halophyte, Atriplex canescens. Sequence analysis has revealed that AcHMA1 contains two heavy metal binding domains. Treatments with metals (Fe, Cu, Ni, Cd or Pb), PEG6000 and NaHCO3 highly induced AcHMA1 expression in A. canescens, whereas NaCl and low temperature decreased its expression. The role of AcHMA1 in metal stress tolerance was examined using a yeast expression system. Expression of the AcHMA1 gene significantly increased the ability of yeast cells to adapt to and recover from exposure to excess iron. AcHMA1 expression also provided salt, alkaline, osmotic and oxidant stress tolerance in yeast cells. Finally, subcellular localization of an AcHMA1/GFP fusion protein expressed in tobacco cells showed that AcHMA1 was localized in the plasma membrane. Thus, our results suggest that AcHMA1 encodes a membrane-localized metal tolerance protein that mediates the detoxification of iron in eukaryotes. Furthermore, AcHMA1 also participates in the response to abiotic stress.

Fates of Fe3O4 and Fe3O4@SiO2 nanoparticles in human mesenchymal stem cells assessed by synchrotron radiation-based techniques.

Superparamagnetic iron oxide nanoparticles (SPIOs) have been widely used as the magnetic resonance imaging (MRI) contrast agent in biomedical studies and clinical applications, with special interest recently in in vivo stem cell tracking. However, a full understanding of the fate of SPIOs in cells has not been achieved yet, which is particularly important for stem cells since any change of the microenvironment may disturb their propagation and differentiation behaviors. Herein, synchrotron radiation-based X-ray fluorescence (XRF) in combination with X-ray absorption spectroscopy (XAS) were used to in situ reveal the fate of Fe3O4 and Fe3O4@SiO2 NPs in human mesenchymal stem cells (hMSCs), in which the dynamic changes of their distribution and chemical speciation were precisely determined. The XAS analysis evidences that Fe3O4 NPs cultured with hMSCs are quite stable and almost keep their initial chemical form up to 14 days, which is contradictory to the previous report that Fe3O4 NPs were unstable in cell labeling assessed by using a simplified lysosomal model system. Coating with a SiO2 shell, Fe3O4@SiO2 NPs present higher stability in hMSCs without detectable changes of their chemical form. In addition, XRF analysis demonstrates that Fe3O4@SiO2 NPs can label hMSCs in a high efficiency manner and are solely distributed in cytoplasm during cell proliferation, making it an ideal probe for in vivo stem cell tracking. These findings with the help of synchrotron radiation-based XAS and XRF improve our understanding of the fate of SPIOs administered to hMSCs and will help the future design of SPIOs for safe and efficient stem cells tracking.