Small RNAs(sRNAs)are essential for regulating plant growth and development,and they possess the notable ability to travel long distances within organisms to regulate target gene expression.Our study examined the dcl2 ...Small RNAs(sRNAs)are essential for regulating plant growth and development,and they possess the notable ability to travel long distances within organisms to regulate target gene expression.Our study examined the dcl2 mutant,a key enzyme in s RNA biogenesis,to determine the role of the DCL2 protein in s RNA synthesis and to identify mobile s RNAs under DCL2 regulation.Through grafting experiments between dcl2 mutants and wild-type soybean plants,coupled with s RNA sequencing,we identified14,105 s RNAs significantly affected by DCL2 and discovered 375 mobile s RNAs under its regulation.Degradome analysis provided deeper insights into the regulatory effects of these mobile s RNAs on their target genes,enabling us to understand their potential influences on plant development and stress responses.Leveraging the systemic movement of s RNAs from roots to shoots,we propose a novel strategy for manipulating gene expression in aboveground tissues.Overall,our research findings not only deepen our understanding of the complex regulatory networks involving mobile s RNAs regulated by DCL2,but also provide a new strategy for gene regulation,which could have a positive impact on agricultural biotechnology.展开更多
BACKGROUND Growth arrest-specific gene 2(GAS2)plays a role in modulating in reversible growth arrest cell cycle,apoptosis,and cell survival.GAS2 protein is universally expressed in most normal tissues,particularly in ...BACKGROUND Growth arrest-specific gene 2(GAS2)plays a role in modulating in reversible growth arrest cell cycle,apoptosis,and cell survival.GAS2 protein is universally expressed in most normal tissues,particularly in the liver,but is depleted in some tumor tissues.However,the functional mechanisms of GAS2 in hepatocellular carcinoma(HCC)are not fully defined.AIM To investigate the function and mechanism of GAS2 in HCC.METHODS GAS2 expression in clinic liver and HCC specimens was analyzed by real-time PCR and western blotting.Cell proliferation was analyzed by counting,MTS,and colony formation assays.Cell cycle analysis was performed by flow cytometry.Cell apoptosis was investigated by Annexin V apoptosis assay and western blotting.RESULTS GAS2 protein expression was lower in HCC than in normal tissues.Overexpression of GAS2 inhibited the proliferation of HCC cells with wide-type p53,while knockdown of GAS2 promoted the proliferation of hepatocytes(P<0.05).Furthermore,GAS2 overexpression impeded the G1-to-S cell cycle transition and arrested more G1 cells,particularly the elevation of sub G1(P<0.01).Apoptosis induced by GAS2 was dependent on p53,which was increased by etoposide addition.The expression of p53 and apoptosis markers was further enhanced when GAS2 was upregulated,but became diminished upon downregulation of GAS2.In the clinic specimen,GAS2 was downregulated in more than 60%of HCCs.The average fold changes of GAS2 expression in tumor tissues were significantly lower than those in paired non-tumor tissues(P<0.05).CONCLUSION GAS2 plays a vital role in HCC cell proliferation and apoptosis,possibly by regulating the cell cycle and p53-dependent apoptosis pathway.展开更多
Periodontitis is a widespread oral disease characterized by continuous inflammation of the periodontal tissue and an irreversible alveolar bone loss, which eventually leads to tooth loss. Four-octyl itaconate(4-OI) is...Periodontitis is a widespread oral disease characterized by continuous inflammation of the periodontal tissue and an irreversible alveolar bone loss, which eventually leads to tooth loss. Four-octyl itaconate(4-OI) is a cell-permeable itaconate derivative and has been recognized as a promising therapeutic target for the treatment of inflammatory diseases. Here, we explored, for the first time,the protective effect of 4-OI on inhibiting periodontal destruction, ameliorating local inflammation, and the underlying mechanism in periodontitis. Here we showed that 4-OI treatment ameliorates inflammation induced by lipopolysaccharide in the periodontal microenvironment. 4-OI can also significantly alleviate inflammation and alveolar bone loss via Nrf2 activation as observed on samples from experimental periodontitis in the C57BL/6 mice. This was further confirmed as silencing Nrf2 blocked the antioxidant effect of 4-OI by downregulating the expression of downstream antioxidant enzymes. Additionally, molecular docking simulation indicated the possible mechanism under Nrf2 activation. Also, in Nrf2-/-mice, 4-OI treatment did not protect against alveolar bone dysfunction due to induced periodontitis, which underlined the importance of the Nrf2 in 4-OI mediated periodontitis treatment.Our results indicated that 4-OI attenuates inflammation and oxidative stress via disassociation of KEAP1-Nrf2 and activation of Nrf2 signaling cascade. Taken together, local administration of 4-OI offers clinical potential to inhibit periodontal destruction,ameliorate local inflammation for more predictable periodontitis.展开更多
The study aims to confirm the neuroregenerative effects of bacterial melanin （BM） on central nervous system injury using a special staining method based on the detection of Ca^2＋-dependent acid phosphatase activity...The study aims to confirm the neuroregenerative effects of bacterial melanin （BM） on central nervous system injury using a special staining method based on the detection of Ca^2＋-dependent acid phosphatase activity. Twenty-four rats were randomly assigned to undergo either unilateral destruction of sensorimotor cortex （group I; n = 12） or unilateral rubrospinal tract transection at the cervical level （C3-4） （group II; n = 12）. In each group, six rats were randomly selected after surgery to undergo intramuscular injection of BM solution （BM subgroup） and the remaining six rats were intramuscularly in）ected with saline （saline subgroup）. Neurological testing confirmed that BM accelerated the recovery of motor function in rats from both BM and saline subgroups. Two months after surgery, Ca^2＋-dependent acid phosphatase activity detection in combination with Chilingarian＇s calcium adenoside triphosphate method revealed that BM stimulated the sprouting of fibers and dilated the capillaries in the brain and spinal cord. These results suggest that BM can promote the recovery of motor function of rats with central nervous system injury; and detection of Ca^2＋-dependent acid phosphatase activity is a fast and easy method used to study the regeneration-promoting effects of BM on the injured central nervous system.展开更多
Tumor microenvironment(TME),as the“soil”of tumor growth and metastasis,exhibits significant differences from normal physiological conditions.However,how to manipulate the distinctions to achieve the accurate therapy...Tumor microenvironment(TME),as the“soil”of tumor growth and metastasis,exhibits significant differences from normal physiological conditions.However,how to manipulate the distinctions to achieve the accurate therapy of primary and metastatic tumors is still a challenge.Herein,an innovative nanoreactor(AH@MBTF)is developed to utilize the apparent differences(copper concentration and H_(2)O_(2)level)between tumor cells and normal cells to eliminate primary tumor based on H_(2)O_(2)-dependent photothermal-chemodynamic therapy and suppress metastatic tumor through copper complexation.This nanoreactor is constructed using functionalized MSN incorporating benzoyl thiourea(BTU),triphenylphosphine(TPP),and folic acid(FA),while being co-loaded with horseradish peroxidase(HRP)and its substrate ABTS.During therapy,the BTU moieties on AH@MBTF could capture excessive copper(highly correlated with tumor metastasis),presenting exceptional anti-metastasis activity.Simultaneously,the complexation between BTU and copper triggers the formation of cuprous ions,which further react with H_(2)O_(2)to generate cytotoxic hydroxyl radical(•OH),inhibiting tumor growth via che-modynamic therapy.Additionally,the stepwise targeting of FA and TPP guides AH@MBTF to accurately accu-mulate in tumor mitochondria,containing abnormally high levels of H_(2)O_(2).As a catalyst,HRP mediates the oxidation reaction between ABTS and H_(2)O_(2)to yield activated ABTS•^(+).Upon 808 nm laser irradiation,the activated ABTS•^(+)performs tumor-specific photothermal therapy,achieving the ablation of primary tumor by raising the tissue temperature.Collectively,this intelligent nanoreactor possesses profound potential in inhib-iting tumor progression and metastasis.展开更多
The vast osteocytic network is believed to orchestrate bone metabolic activity in response to mechanical stimuli through production of sclerostin, RANKL, and osteoprotegerin(OPG). However, the mechanisms of osteocyte ...The vast osteocytic network is believed to orchestrate bone metabolic activity in response to mechanical stimuli through production of sclerostin, RANKL, and osteoprotegerin(OPG). However, the mechanisms of osteocyte mechanotransduction remain poorly understood. We've previously shown that osteocyte mechanosensitivity is encoded through unique intracellular calcium (Ca^(2+) ) dynamics. Here, by simultaneously monitoring Ca^(2+) and actin dynamics in single cells exposed to fluid shear flow, we detected actin network contractions immediately upon onset of flow-induced Ca^(2+) transients, which were facilitated by smooth muscle myosin and further confirmed in native osteocytes ex vivo. Actomyosin contractions have been linked to the secretion of extracellular vesicles(EVs), and our studies demonstrate that mechanical stimulation upregulates EV production in osteocytes through immunostaining for the secretory vesicle marker Lysosomal-associated membrane protein 1(LAMP1) and quantifying EV release in conditioned medium, both of which are blunted when Ca^(2+) signaling was inhibited by neomycin. Axial tibia compression was used to induce anabolic bone formation responses in mice, revealing upregulated LAMP1 and expected downregulation of sclerostin in vivo. This load-related increase in LAMP1 expression was inhibited in neomycin-injected mice compared to vehicle.Micro-computed tomography revealed significant load-related increases in both trabecular bone volume fraction and cortical thickness after two weeks of loading, which were blunted by neomycin treatment. In summary, we found mechanical stimulation of osteocytes activates Ca^(2+) -dependent contractions and enhances the production and release of EVs containing bone regulatory proteins. Further, blocking Ca^(2+) signaling significantly attenuates adaptation to mechanical loading in vivo, suggesting a critical role for Ca^(2+) -mediated signaling in bone adaptation.展开更多
Objective To test whether in the absence of actin, actin-binding proteins such as caldesmon, calponin, and tropomyosin interact with the myosin of unphosphorylation, Ca 2+ -dependent phosphorylation (CDP), and Ca 2+ -...Objective To test whether in the absence of actin, actin-binding proteins such as caldesmon, calponin, and tropomyosin interact with the myosin of unphosphorylation, Ca 2+ -dependent phosphorylation (CDP), and Ca 2+ -independent phosphorylati-on (CIP) and stimulate myosin Mg 2+ -ATPase activities. Methods Mg 2+ -ATPase activities were measured to evaluate the effects of caldesmon, calponin, and tropomyosin on the myosin in unphosphorylation, CDP by myosin light chain kinase (MLCK), and CIP by MLCK. Results (1) At different incubation-time, i.e., 5, 10, 20, 40, and 60 minutes, the highest Mg 2+ -ATPase activity was ob-served when myosin was in the state of CDP, the medium was CIP of myosin, and the lowest was the unphosphorylated myosin. (2) In the absence of caldesmon, calponin, and tropomyosin, the Mg 2+ -ATPase activities from high to low were in the following order: CDP, CIP, and unphosphorylated myosin. However, in the presence of caldesmon, calponin, and tropo-myosin, the order of relative value of Mg 2+ -ATPase activities from high to low was unphosphorylated, CIP, and CDP of myosin respectively compared to the corresponding controls. Conclusions The results propose that caldesmon, calponin, and tropomyosin are capable of stimulating Mg 2+ -ATPase activity of smooth muscle myosin in Ca 2+ -independent manner, since Ca 2+ is not obligating for the stimulating effects of the three proteins. The common characteristic of the three proteins is that when myosin activities are low, their activations are relatively strong and this property might be involved in smooth muscle tension keeping.展开更多
Transcription factors can be used to engineer plants for enhanced productivity.However,the mechanism(s)by which the C2H2-type zinc fi nger transcription factor enhances pathogen resistance in cells is not fully unders...Transcription factors can be used to engineer plants for enhanced productivity.However,the mechanism(s)by which the C2H2-type zinc fi nger transcription factor enhances pathogen resistance in cells is not fully understood.Here,Agrobacterium tumefaciens carrying the gene for Arabidopsis thaliana cysteine2/histidine2-type transcription factor 6(ZAT6)was used to engineer rice(Oryza sativa L.),cotton(Gossypium hirsutum L.),and slash pine(Pinus elliottii Engelm.)to generate transgenic cell lines.Transgenic cells were then inoculated with the pathogenic bacterium Pseudomonas syringae.Compared to the control,cell viability of transgenic cells increased 39–47%and growth rate increased 9–15%by 7 days after inoculation in rice,cotton and slash pine.Acid phosphatase activity and alkaline phosphatase activity and transcript levels of Ca 2+-dependent protein kinase genes OsCPK1,OsCPK2,OsCPK6,and OsCPK8 and mitogen-activated protein kinase genes OsMAPK1,OsMAPK2,OsMAPK3,and OsMAPK8 increased signifi cantly in transgenic rice cells by 3 day after inoculation,and extracellular pH had decreased by 10–14%by 96 min after inoculation in transgenic rice,cotton and slash pine cells.These results suggest that ZAT6 enhances P.syringae resistance in plant cells by modulating transcription of CPK and MAPK and oxidase activity.展开更多
Secondary xylem development has long been recognized as a typical case of programmed cell death （PCD） in plants. During PCD, the degradation of genomic DNA is catalyzed by endonucleases. However, to date, no endonuc...Secondary xylem development has long been recognized as a typical case of programmed cell death （PCD） in plants. During PCD, the degradation of genomic DNA is catalyzed by endonucleases. However, to date, no endonuclease has been shown to participate in secondary xylem development. Two novel Ca^2＋-dependent DNase genes, EuCaN1 and EuCaN2, were identified from the differentiating secondary xylem of the tree Eucommia ulmoides Oliv., their functions were studied by DNase activity assay, in situ hybridization, protein immunolocalization and virus-induced gene silencing experiments. Full-length cDNAs of EuCaN1 and EuCaN2 contained an open reading frame of 987 bp, encoding two proteins of 328 amino acids with SNase-like functional domains. The genomic DNA sequence for EuCaN1 had no introns, while EuCaN2 had 8 introns. EuCaN1 and EuCaN2 digested ssDNA and dsDNA with Ca^2＋-dependence at neutral pH. Their expression was confined to differentiating secondary xylem cells and the proteins were localized in the nucleus. Their activity dynamics was closely correlated with secondary xylem development. Secondary xylem cell differentiation is influenced by RNAi of endonuclease genes. The results provide evidence that the Ca^2＋-dependent DNases are involved in secondary xylem development.展开更多
Viruses can infect host plants to cause severe diseases and substantial agricultural loss, while plants have evolved RNA interference （RNAi） strategy to defend against viral infection. Despite enormous efforts, only...Viruses can infect host plants to cause severe diseases and substantial agricultural loss, while plants have evolved RNA interference （RNAi） strategy to defend against viral infection. Despite enormous efforts, only a few host proteins in RNAi pathway were shown to mediate antiviral defense, including RNA-dependent RNA polymerase I （RDRI）, RDR6, DICER-LIKE 2 （DCL2） and DCL4. In this study, we carried out a genetic screen for antiviral factors of RNAi pathway in Arabidopsis rdr6 background via inoculation with a 2b- deficient Cucumber Mosaic Virus （CMV-△2b）. We identified a mutant susceptible to CMV-△2h, referred to as enhancer o ojrdr6 （enor） 3-1 rdr6, and found that ENOR3 encodes a functionally unknown protein with high homology to the mammalian Non Imprinted in Prader-Willi/Angelman （NIPA） magnesium transporters. ENOR3 inhibits accumulation of CMV-△2b and acts additively with RDR1, RDR6, DCL2 and DCL4 in antivira/ defense. These results uncover that ENOR3 is a key component in antiviral RNAi Dathwav, and provide new insights into antiviral immunity.展开更多
Many environmental chemicals and pesticides have been found to alter neuroendocrine communication in exposed biological objects. The environmental loads have primary and secondary effects that can alter the homeostati...Many environmental chemicals and pesticides have been found to alter neuroendocrine communication in exposed biological objects. The environmental loads have primary and secondary effects that can alter the homeostatic regulation potential. Since it is difficult to avoid human exposition, a potentially important area of research to develop in vivo and in vitro experimental models. In this context, the primary aim of this study was to demonstrate the effects of chlorobenzenes on adrenocorticotrophic hormone（ACTH） release. In our experimental study, male Wistar rats were exposed to 0.1, 1.0 and 10 μg/b.w.（body weight） kg of 1,2,4-trichlorobenzene and hexachlorobenzene（Cl B） mix via gastric tube for 30, 60 or 90 days. At the endpoints of the experiment blood samples were taken and animals were decapitated. Primary,monolayer adenohypophysis cell cultures were prepared by enzymatic and mechanical digestion. The ACTH hormone content in serum and supernatant media was measured by immuno-chemiluminescence assay. The Mg2＋-dependent ATPase activity was determined by modified method of Martin and Dotty. Significant differences were detected in the hormone release between the control and treated groups. The hormone release was enhanced characteristically in exposed groups depending upon the dose and duration of exposure. The Mg2＋-ATPase activity enhanced after chronic and subtoxic Cl B exposition. Light microscopy revealed that the adenohypophysis seemed to be more abundant. Results indicate that Wistar rats exposed to subtoxic Cl B have direct and indirect effects on hypothalamus–hypophysis–adrenal axis.展开更多
AIM To filtrate breast cancer resistance protein(BCRP)-mediated resistance agents and investigatethe mechanism,so as to provide valuable datum for optimization clinical chemotherapy scheme to tumor withevaluation mark...AIM To filtrate breast cancer resistance protein(BCRP)-mediated resistance agents and investigatethe mechanism,so as to provide valuable datum for optimization clinical chemotherapy scheme to tumor withevaluation marker of BCRP expression.METHODS MTT assay was used to filtrate BCRP-mediatedresistance agents with PA317/Tet-on/TRE-BCRP cell of different expression levels of BCRP after treated withdifferent concentration anticancer agents.High performance liquid chromatography(HPLC) was applied tomeasure relative dose of intracellular retention resistance agents.Nuclear DNA fluorescence dye,Hochest33258,staining and flow cytometry were adopted to detect apoptotic cells after treated with drugs.RESULTSThere were shown increasing durg-resistance to 5-fluorouracil,methotrexate,doxirubicin,pirarubicin,etoposide and mitoxantrone followed with increasing expression of BCRP on PA317/Tet-on/TRE-BCRPcells(P<0.05,n=3),but shown sensitive to paclitaxel,cisplatin,vincristine,mitomycin and vindesine.Therealso was shown significant negative correlation between the intracellular retention dose of 5-fluorouracil withdifferent expression of BCRP(r=-0.885,P<0.05,n=3).There were shown parallel results ofthat decreasingcellular apoptotic rate with increasing cellular expression of BCRP after treated with 5-fluorouracil byfluorescence dye staining and flow cytometry(P<0.05,n=3),and also shown significate rise of the apoptoticrate of BCRP expression cells after treated with Ko143 (P<0.05,n=3).Every group of cells could be differentextently blocked in phase of G_0/G_1 treated with 5-fluorouracil.CONCLUSION Resistance of 5-fluorouracilcould be especially mediated by conjugated with BCRP and acted as drug exclude-pump substrate.Cellularability resistant to 5-fluorouracil-induced apoptosis could be reinforced by BCRP expression.展开更多
This brief article highlights the results of Fu et al.(Proc Natl Acad Sci USA 119:e2204574119,2022),who recently found that manganese(Mn)deficiency triggers long-lasting multicellular Ca^(2+) oscillations in the elong...This brief article highlights the results of Fu et al.(Proc Natl Acad Sci USA 119:e2204574119,2022),who recently found that manganese(Mn)deficiency triggers long-lasting multicellular Ca^(2+) oscillations in the elongation zone(EZ)of Arabidopsis roots and revealed a Ca^(2+)-CPK21/23-NRAMP1 axis as an important mechanism for plant tolerance and adaptation to low Mn.展开更多
基金supported by the Strategic Priority Research Program of the Chinese Academy of Sciences(XDA24010205)the CAS Project for Young Scientists in Basic Research(YSBR-011)+1 种基金National Natural Science Foundation of China(32272101)the Agricultural Science and Technology Innovation Program of CAAS。
文摘Small RNAs(sRNAs)are essential for regulating plant growth and development,and they possess the notable ability to travel long distances within organisms to regulate target gene expression.Our study examined the dcl2 mutant,a key enzyme in s RNA biogenesis,to determine the role of the DCL2 protein in s RNA synthesis and to identify mobile s RNAs under DCL2 regulation.Through grafting experiments between dcl2 mutants and wild-type soybean plants,coupled with s RNA sequencing,we identified14,105 s RNAs significantly affected by DCL2 and discovered 375 mobile s RNAs under its regulation.Degradome analysis provided deeper insights into the regulatory effects of these mobile s RNAs on their target genes,enabling us to understand their potential influences on plant development and stress responses.Leveraging the systemic movement of s RNAs from roots to shoots,we propose a novel strategy for manipulating gene expression in aboveground tissues.Overall,our research findings not only deepen our understanding of the complex regulatory networks involving mobile s RNAs regulated by DCL2,but also provide a new strategy for gene regulation,which could have a positive impact on agricultural biotechnology.
基金Supported by the National Natural Science Foundation of China,No.81702777Natural Science Foundation of Guangdong Province,No.2015A030310053
文摘BACKGROUND Growth arrest-specific gene 2(GAS2)plays a role in modulating in reversible growth arrest cell cycle,apoptosis,and cell survival.GAS2 protein is universally expressed in most normal tissues,particularly in the liver,but is depleted in some tumor tissues.However,the functional mechanisms of GAS2 in hepatocellular carcinoma(HCC)are not fully defined.AIM To investigate the function and mechanism of GAS2 in HCC.METHODS GAS2 expression in clinic liver and HCC specimens was analyzed by real-time PCR and western blotting.Cell proliferation was analyzed by counting,MTS,and colony formation assays.Cell cycle analysis was performed by flow cytometry.Cell apoptosis was investigated by Annexin V apoptosis assay and western blotting.RESULTS GAS2 protein expression was lower in HCC than in normal tissues.Overexpression of GAS2 inhibited the proliferation of HCC cells with wide-type p53,while knockdown of GAS2 promoted the proliferation of hepatocytes(P<0.05).Furthermore,GAS2 overexpression impeded the G1-to-S cell cycle transition and arrested more G1 cells,particularly the elevation of sub G1(P<0.01).Apoptosis induced by GAS2 was dependent on p53,which was increased by etoposide addition.The expression of p53 and apoptosis markers was further enhanced when GAS2 was upregulated,but became diminished upon downregulation of GAS2.In the clinic specimen,GAS2 was downregulated in more than 60%of HCCs.The average fold changes of GAS2 expression in tumor tissues were significantly lower than those in paired non-tumor tissues(P<0.05).CONCLUSION GAS2 plays a vital role in HCC cell proliferation and apoptosis,possibly by regulating the cell cycle and p53-dependent apoptosis pathway.
基金supported by the National Natural Science Foundation of China (31971282 and 32071362)2019 Chongqing Graduate Tutor Team Construction Project (dstd201903)Scientific and Technological Research Program of Chongqing Municipal Education Commission (KJQN201900415)。
文摘Periodontitis is a widespread oral disease characterized by continuous inflammation of the periodontal tissue and an irreversible alveolar bone loss, which eventually leads to tooth loss. Four-octyl itaconate(4-OI) is a cell-permeable itaconate derivative and has been recognized as a promising therapeutic target for the treatment of inflammatory diseases. Here, we explored, for the first time,the protective effect of 4-OI on inhibiting periodontal destruction, ameliorating local inflammation, and the underlying mechanism in periodontitis. Here we showed that 4-OI treatment ameliorates inflammation induced by lipopolysaccharide in the periodontal microenvironment. 4-OI can also significantly alleviate inflammation and alveolar bone loss via Nrf2 activation as observed on samples from experimental periodontitis in the C57BL/6 mice. This was further confirmed as silencing Nrf2 blocked the antioxidant effect of 4-OI by downregulating the expression of downstream antioxidant enzymes. Additionally, molecular docking simulation indicated the possible mechanism under Nrf2 activation. Also, in Nrf2-/-mice, 4-OI treatment did not protect against alveolar bone dysfunction due to induced periodontitis, which underlined the importance of the Nrf2 in 4-OI mediated periodontitis treatment.Our results indicated that 4-OI attenuates inflammation and oxidative stress via disassociation of KEAP1-Nrf2 and activation of Nrf2 signaling cascade. Taken together, local administration of 4-OI offers clinical potential to inhibit periodontal destruction,ameliorate local inflammation for more predictable periodontitis.
基金supported by the Armenian National Science and Education Fund for Project in New York,USA(No.ANSEF biotech-4241)
文摘The study aims to confirm the neuroregenerative effects of bacterial melanin （BM） on central nervous system injury using a special staining method based on the detection of Ca^2＋-dependent acid phosphatase activity. Twenty-four rats were randomly assigned to undergo either unilateral destruction of sensorimotor cortex （group I; n = 12） or unilateral rubrospinal tract transection at the cervical level （C3-4） （group II; n = 12）. In each group, six rats were randomly selected after surgery to undergo intramuscular injection of BM solution （BM subgroup） and the remaining six rats were intramuscularly in）ected with saline （saline subgroup）. Neurological testing confirmed that BM accelerated the recovery of motor function in rats from both BM and saline subgroups. Two months after surgery, Ca^2＋-dependent acid phosphatase activity detection in combination with Chilingarian＇s calcium adenoside triphosphate method revealed that BM stimulated the sprouting of fibers and dilated the capillaries in the brain and spinal cord. These results suggest that BM can promote the recovery of motor function of rats with central nervous system injury; and detection of Ca^2＋-dependent acid phosphatase activity is a fast and easy method used to study the regeneration-promoting effects of BM on the injured central nervous system.
基金supported by the National High Level Talents Special Support Plan(X.C.)the National Natural Science Foundation of China(82272141 to X.C.)+4 种基金the“Young Talent Support Plan”of Xi’an Jiaotong University(X.C.)the Shaanxi Innovative Research Team of Science and Technology(S2023-ZC-TD-0152)the Natural Science Foundation of Shaanxi Province(2022JZ-48 to X.C.)the National Key Research and Development Program of China(2023YFC2509104 to X.C.)the Postdoctoral Science Foundation of China(2023M732812 to T.L.).
文摘Tumor microenvironment(TME),as the“soil”of tumor growth and metastasis,exhibits significant differences from normal physiological conditions.However,how to manipulate the distinctions to achieve the accurate therapy of primary and metastatic tumors is still a challenge.Herein,an innovative nanoreactor(AH@MBTF)is developed to utilize the apparent differences(copper concentration and H_(2)O_(2)level)between tumor cells and normal cells to eliminate primary tumor based on H_(2)O_(2)-dependent photothermal-chemodynamic therapy and suppress metastatic tumor through copper complexation.This nanoreactor is constructed using functionalized MSN incorporating benzoyl thiourea(BTU),triphenylphosphine(TPP),and folic acid(FA),while being co-loaded with horseradish peroxidase(HRP)and its substrate ABTS.During therapy,the BTU moieties on AH@MBTF could capture excessive copper(highly correlated with tumor metastasis),presenting exceptional anti-metastasis activity.Simultaneously,the complexation between BTU and copper triggers the formation of cuprous ions,which further react with H_(2)O_(2)to generate cytotoxic hydroxyl radical(•OH),inhibiting tumor growth via che-modynamic therapy.Additionally,the stepwise targeting of FA and TPP guides AH@MBTF to accurately accu-mulate in tumor mitochondria,containing abnormally high levels of H_(2)O_(2).As a catalyst,HRP mediates the oxidation reaction between ABTS and H_(2)O_(2)to yield activated ABTS•^(+).Upon 808 nm laser irradiation,the activated ABTS•^(+)performs tumor-specific photothermal therapy,achieving the ablation of primary tumor by raising the tissue temperature.Collectively,this intelligent nanoreactor possesses profound potential in inhib-iting tumor progression and metastasis.
基金supported by NIH R01 AR052461 and NIH R01 AR069148supported by a NSF Graduate Research Fellowship. A. E. M.supported by training grant T32 AR059038
文摘The vast osteocytic network is believed to orchestrate bone metabolic activity in response to mechanical stimuli through production of sclerostin, RANKL, and osteoprotegerin(OPG). However, the mechanisms of osteocyte mechanotransduction remain poorly understood. We've previously shown that osteocyte mechanosensitivity is encoded through unique intracellular calcium (Ca^(2+) ) dynamics. Here, by simultaneously monitoring Ca^(2+) and actin dynamics in single cells exposed to fluid shear flow, we detected actin network contractions immediately upon onset of flow-induced Ca^(2+) transients, which were facilitated by smooth muscle myosin and further confirmed in native osteocytes ex vivo. Actomyosin contractions have been linked to the secretion of extracellular vesicles(EVs), and our studies demonstrate that mechanical stimulation upregulates EV production in osteocytes through immunostaining for the secretory vesicle marker Lysosomal-associated membrane protein 1(LAMP1) and quantifying EV release in conditioned medium, both of which are blunted when Ca^(2+) signaling was inhibited by neomycin. Axial tibia compression was used to induce anabolic bone formation responses in mice, revealing upregulated LAMP1 and expected downregulation of sclerostin in vivo. This load-related increase in LAMP1 expression was inhibited in neomycin-injected mice compared to vehicle.Micro-computed tomography revealed significant load-related increases in both trabecular bone volume fraction and cortical thickness after two weeks of loading, which were blunted by neomycin treatment. In summary, we found mechanical stimulation of osteocytes activates Ca^(2+) -dependent contractions and enhances the production and release of EVs containing bone regulatory proteins. Further, blocking Ca^(2+) signaling significantly attenuates adaptation to mechanical loading in vivo, suggesting a critical role for Ca^(2+) -mediated signaling in bone adaptation.
基金Supported by the National Natural Science Foundation of China ( 30070203).
文摘Objective To test whether in the absence of actin, actin-binding proteins such as caldesmon, calponin, and tropomyosin interact with the myosin of unphosphorylation, Ca 2+ -dependent phosphorylation (CDP), and Ca 2+ -independent phosphorylati-on (CIP) and stimulate myosin Mg 2+ -ATPase activities. Methods Mg 2+ -ATPase activities were measured to evaluate the effects of caldesmon, calponin, and tropomyosin on the myosin in unphosphorylation, CDP by myosin light chain kinase (MLCK), and CIP by MLCK. Results (1) At different incubation-time, i.e., 5, 10, 20, 40, and 60 minutes, the highest Mg 2+ -ATPase activity was ob-served when myosin was in the state of CDP, the medium was CIP of myosin, and the lowest was the unphosphorylated myosin. (2) In the absence of caldesmon, calponin, and tropomyosin, the Mg 2+ -ATPase activities from high to low were in the following order: CDP, CIP, and unphosphorylated myosin. However, in the presence of caldesmon, calponin, and tropo-myosin, the order of relative value of Mg 2+ -ATPase activities from high to low was unphosphorylated, CIP, and CDP of myosin respectively compared to the corresponding controls. Conclusions The results propose that caldesmon, calponin, and tropomyosin are capable of stimulating Mg 2+ -ATPase activity of smooth muscle myosin in Ca 2+ -independent manner, since Ca 2+ is not obligating for the stimulating effects of the three proteins. The common characteristic of the three proteins is that when myosin activities are low, their activations are relatively strong and this property might be involved in smooth muscle tension keeping.
文摘Transcription factors can be used to engineer plants for enhanced productivity.However,the mechanism(s)by which the C2H2-type zinc fi nger transcription factor enhances pathogen resistance in cells is not fully understood.Here,Agrobacterium tumefaciens carrying the gene for Arabidopsis thaliana cysteine2/histidine2-type transcription factor 6(ZAT6)was used to engineer rice(Oryza sativa L.),cotton(Gossypium hirsutum L.),and slash pine(Pinus elliottii Engelm.)to generate transgenic cell lines.Transgenic cells were then inoculated with the pathogenic bacterium Pseudomonas syringae.Compared to the control,cell viability of transgenic cells increased 39–47%and growth rate increased 9–15%by 7 days after inoculation in rice,cotton and slash pine.Acid phosphatase activity and alkaline phosphatase activity and transcript levels of Ca 2+-dependent protein kinase genes OsCPK1,OsCPK2,OsCPK6,and OsCPK8 and mitogen-activated protein kinase genes OsMAPK1,OsMAPK2,OsMAPK3,and OsMAPK8 increased signifi cantly in transgenic rice cells by 3 day after inoculation,and extracellular pH had decreased by 10–14%by 96 min after inoculation in transgenic rice,cotton and slash pine cells.These results suggest that ZAT6 enhances P.syringae resistance in plant cells by modulating transcription of CPK and MAPK and oxidase activity.
基金supported by the National Basic Research Program of China (2012CB114500)the National Natural Science Foundation of China (31070156)
文摘Secondary xylem development has long been recognized as a typical case of programmed cell death （PCD） in plants. During PCD, the degradation of genomic DNA is catalyzed by endonucleases. However, to date, no endonuclease has been shown to participate in secondary xylem development. Two novel Ca^2＋-dependent DNase genes, EuCaN1 and EuCaN2, were identified from the differentiating secondary xylem of the tree Eucommia ulmoides Oliv., their functions were studied by DNase activity assay, in situ hybridization, protein immunolocalization and virus-induced gene silencing experiments. Full-length cDNAs of EuCaN1 and EuCaN2 contained an open reading frame of 987 bp, encoding two proteins of 328 amino acids with SNase-like functional domains. The genomic DNA sequence for EuCaN1 had no introns, while EuCaN2 had 8 introns. EuCaN1 and EuCaN2 digested ssDNA and dsDNA with Ca^2＋-dependence at neutral pH. Their expression was confined to differentiating secondary xylem cells and the proteins were localized in the nucleus. Their activity dynamics was closely correlated with secondary xylem development. Secondary xylem cell differentiation is influenced by RNAi of endonuclease genes. The results provide evidence that the Ca^2＋-dependent DNases are involved in secondary xylem development.
基金financially supported by the National Natural Science Foundation of China (Nos. 31421001 and 31630085)the National Key R&D Program of China (2016YFA0500501)
文摘Viruses can infect host plants to cause severe diseases and substantial agricultural loss, while plants have evolved RNA interference （RNAi） strategy to defend against viral infection. Despite enormous efforts, only a few host proteins in RNAi pathway were shown to mediate antiviral defense, including RNA-dependent RNA polymerase I （RDRI）, RDR6, DICER-LIKE 2 （DCL2） and DCL4. In this study, we carried out a genetic screen for antiviral factors of RNAi pathway in Arabidopsis rdr6 background via inoculation with a 2b- deficient Cucumber Mosaic Virus （CMV-△2b）. We identified a mutant susceptible to CMV-△2h, referred to as enhancer o ojrdr6 （enor） 3-1 rdr6, and found that ENOR3 encodes a functionally unknown protein with high homology to the mammalian Non Imprinted in Prader-Willi/Angelman （NIPA） magnesium transporters. ENOR3 inhibits accumulation of CMV-△2b and acts additively with RDR1, RDR6, DCL2 and DCL4 in antivira/ defense. These results uncover that ENOR3 is a key component in antiviral RNAi Dathwav, and provide new insights into antiviral immunity.
基金supported by TáMOP - 4.2.2.A-11/1/KONV - 2012-0047supported by the European Union and the State of Hungary+1 种基金co-financed by the European Social Fund in the framework of TáMOP 4.2.4.A/2-11-1-2012-0001 ‘ National Excellence Program ’supported by TáMOP - 4.1.1.C-12/1/KONV - 2012-0012
文摘Many environmental chemicals and pesticides have been found to alter neuroendocrine communication in exposed biological objects. The environmental loads have primary and secondary effects that can alter the homeostatic regulation potential. Since it is difficult to avoid human exposition, a potentially important area of research to develop in vivo and in vitro experimental models. In this context, the primary aim of this study was to demonstrate the effects of chlorobenzenes on adrenocorticotrophic hormone（ACTH） release. In our experimental study, male Wistar rats were exposed to 0.1, 1.0 and 10 μg/b.w.（body weight） kg of 1,2,4-trichlorobenzene and hexachlorobenzene（Cl B） mix via gastric tube for 30, 60 or 90 days. At the endpoints of the experiment blood samples were taken and animals were decapitated. Primary,monolayer adenohypophysis cell cultures were prepared by enzymatic and mechanical digestion. The ACTH hormone content in serum and supernatant media was measured by immuno-chemiluminescence assay. The Mg2＋-dependent ATPase activity was determined by modified method of Martin and Dotty. Significant differences were detected in the hormone release between the control and treated groups. The hormone release was enhanced characteristically in exposed groups depending upon the dose and duration of exposure. The Mg2＋-ATPase activity enhanced after chronic and subtoxic Cl B exposition. Light microscopy revealed that the adenohypophysis seemed to be more abundant. Results indicate that Wistar rats exposed to subtoxic Cl B have direct and indirect effects on hypothalamus–hypophysis–adrenal axis.
文摘AIM To filtrate breast cancer resistance protein(BCRP)-mediated resistance agents and investigatethe mechanism,so as to provide valuable datum for optimization clinical chemotherapy scheme to tumor withevaluation marker of BCRP expression.METHODS MTT assay was used to filtrate BCRP-mediatedresistance agents with PA317/Tet-on/TRE-BCRP cell of different expression levels of BCRP after treated withdifferent concentration anticancer agents.High performance liquid chromatography(HPLC) was applied tomeasure relative dose of intracellular retention resistance agents.Nuclear DNA fluorescence dye,Hochest33258,staining and flow cytometry were adopted to detect apoptotic cells after treated with drugs.RESULTSThere were shown increasing durg-resistance to 5-fluorouracil,methotrexate,doxirubicin,pirarubicin,etoposide and mitoxantrone followed with increasing expression of BCRP on PA317/Tet-on/TRE-BCRPcells(P<0.05,n=3),but shown sensitive to paclitaxel,cisplatin,vincristine,mitomycin and vindesine.Therealso was shown significant negative correlation between the intracellular retention dose of 5-fluorouracil withdifferent expression of BCRP(r=-0.885,P<0.05,n=3).There were shown parallel results ofthat decreasingcellular apoptotic rate with increasing cellular expression of BCRP after treated with 5-fluorouracil byfluorescence dye staining and flow cytometry(P<0.05,n=3),and also shown significate rise of the apoptoticrate of BCRP expression cells after treated with Ko143 (P<0.05,n=3).Every group of cells could be differentextently blocked in phase of G_0/G_1 treated with 5-fluorouracil.CONCLUSION Resistance of 5-fluorouracilcould be especially mediated by conjugated with BCRP and acted as drug exclude-pump substrate.Cellularability resistant to 5-fluorouracil-induced apoptosis could be reinforced by BCRP expression.
基金supported by grants from National Natural Science Foundation of China(Grant No.31900216)National Key Laboratory of Plant Molecular Genetics.
文摘This brief article highlights the results of Fu et al.(Proc Natl Acad Sci USA 119:e2204574119,2022),who recently found that manganese(Mn)deficiency triggers long-lasting multicellular Ca^(2+) oscillations in the elongation zone(EZ)of Arabidopsis roots and revealed a Ca^(2+)-CPK21/23-NRAMP1 axis as an important mechanism for plant tolerance and adaptation to low Mn.
