AIM： To study the relationship between interleukin-lbeta （IL-1β） up-regulating tissue inhibitor of matrix metalloproteinase-1 （TIMMP-1） mRNA expression and phosphorylation of both c-jun N-terminal kinase （INK）...AIM： To study the relationship between interleukin-lbeta （IL-1β） up-regulating tissue inhibitor of matrix metalloproteinase-1 （TIMMP-1） mRNA expression and phosphorylation of both c-jun N-terminal kinase （INK） and p38 in rat heffatic stellate cells （HSC）. METHODS： RT-PCR was performed to measure the expression of TIMMP-1 mRNA in rat HSC. Western blot was performed to measure IL-1β-induced JNK and p38 activities in rat HSC. RESULTS： TIMMP-1 mRNA expression （1.191± 0.079） was much higher after treatment with IL-1β （10 ng/mL） for 24 h than in control group （0.545±0.091） （P〈0.01）. IL-1β activated INK and p38 in a time-dependent manner. After stimulation with IL-1β for 0, 5, 15, 30, 60 and 120 min, the INK activity was 0.982±0.299, 1.501±0.720, 2.133±0.882, 3.360±0.452, 2.181±0.789, and 1.385 ± 0.368, respectively. There was a significant difference in JNK activity at 15 min （P〈 0.01）, 30 min （P〈 0.01） and 60 min （P〈0.01） in comparison to that at 0 min. The p38 activity was 1.061±0.310, 2.050±0.863, 2.380±0.573, 2.973±0.953, 2.421±0.793, and 1.755 ± 0.433 at the 6 time points （0, 5, 15, 30, 60 and 120 min） respectively. There was a significant difference in p38 activity at 5 min （P〈0.05）, 15 min （P〈0.01）, 30 min （P〈0.01） and 60 min （P〈0.01） compared to that at 0 min. TIMMP-1 mRNA expression trended to decrease in 3 groups pretreated with different concentrations of SP600125 （10 μmol/L, 1.022±0.113; 20 μmol/L, 0.869±0.070; 40 μmol/L, 0.666±0.123）. Their decreases were all significant （P〈0.05, P〈0.01, P〈0.01） in comparison to control group （without SP600125 treatment, 1.163±0.107）. In the other 3 groups pretreated with different concentrations of SB203580 （10 μmol/L, 1.507±0.099; 20 μmol/L, 1.698±0.107; 40 μmol/L, 1.857±0.054）, the expression of TIMMP-1 mRNA increased. Their levels were higher than those in the control group （without SB203580 treatment, 1.027 ± 0.061） with a significant statistical significance （P〈 0.01）. CONCLUSION： IL-1β has a direct action on hepatic fibrosis by up-regulating TIMMP-1 mRNA expression in ratessionin in rate HSC.JNK and p38 mitogen-activated protein kinases （MAPKs） are involved in IL-1β-induced TIMMP-1 gene expression, and play a distinct role in this process, indicating that p38 and .INK pathways cooperatively mediate TIMP-1 mRNA expression in rat HSC.展开更多
Our previous studies showed a predominance of high molecular weight protein group in tumor nuclear matrices. Contrary to normal cells, proteins of this group are preferentially phosphorylated. Phosphoproteins of hepat...Our previous studies showed a predominance of high molecular weight protein group in tumor nuclear matrices. Contrary to normal cells, proteins of this group are preferentially phosphorylated. Phosphoproteins of hepatoma nuclear matrix are selectively subjected to rapid proteolysis. By alkali treatment and a monoclonal antibody against phosphotyrosyl residue the presence of two high molecular weight bands of phosphotyrosyl-containing proteins was detected in nuclear matrices of tumor but not of normal liver cells. High molecular weight protein group of tumor nuclear matrices revealed also a rapid turnover and preferential incorporation of labeled amino acids selectively inhibited by chloramphenicol.展开更多
BACKGROUND Intestinal ischemia reperfusion(I/R)occurs in various diseases,such as trauma and intestinal transplantation.Excessive reactive oxygen species(ROS)accumulation and subsequent apoptotic cell death in intesti...BACKGROUND Intestinal ischemia reperfusion(I/R)occurs in various diseases,such as trauma and intestinal transplantation.Excessive reactive oxygen species(ROS)accumulation and subsequent apoptotic cell death in intestinal epithelia are important causes of I/R injury.PTEN-induced putative kinase 1(PINK1)and phosphorylation of dynamin-related protein 1(DRP1)are critical regulators of ROS and apoptosis.However,the correlation of PINK1 and DRP1 and their function in intestinal I/R injury have not been investigated.Thus,examining the PINK1/DRP1 pathway may help to identify a protective strategy and improve the patient prognosis.AIM To clarify the mechanism of the PINK1/DRP1 pathway in intestinal I/R injury.METHODS Male C57BL/6 mice were used to generate an intestinal I/R model via superior mesenteric artery occlusion followed by reperfusion.Chiu’s score was used to evaluate intestinal mucosa damage.The mitochondrial fission inhibitor mdivi-1 was administered by intraperitoneal injection.Caco-2 cells were incubated in vitro in hypoxia/reoxygenation conditions.Small interfering RNAs and overexpression plasmids were transfected to regulate PINK1 expression.The protein expression levels of PINK1,DRP1,p-DRP1 and cleaved caspase 3 were measured by Western blotting.Cell viability was evaluated using a Cell Counting Kit-8 assay and cell apoptosis was analyzed by TUNEL staining.Mitochondrial fission and ROS were tested by MitoTracker and MitoSOX respectively.RESULTS Intestinal I/R and Caco-2 cell hypoxia/reoxygenation decreased the expression of PINK1 and p-DRP1 Ser637.Pretreatment with mdivi-1 inhibited mitochondrial fission,ROS generation,and apoptosis and ameliorated cell injury in intestinal I/R.Upon PINK1 knockdown or overexpression in vitro,we found that p-DRP1 Ser637 expression and DRP1 recruitment to the mitochondria were associated with PINK1.Furthermore,we verified the physical combination of PINK1 and p-DRP1 Ser637.CONCLUSION PINK1 is correlated with mitochondrial fission and apoptosis by regulating DRP1 phosphorylation in intestinal I/R.These results suggest that the PINK1/DRP1 pathway is involved in intestinal I/R injury,and provide a new approach for prevention and treatment.展开更多
Active and passive anti-Aβimmunotherapies have successfully been used for the prevention and treatment of Alzheimer’s disease animal models.However,clinical use of these immunotherapies is not effective,because the ...Active and passive anti-Aβimmunotherapies have successfully been used for the prevention and treatment of Alzheimer’s disease animal models.However,clinical use of these immunotherapies is not effective,because the vaccination is administered too late.At 1 month of age,100μL of Aβ3–10-KLH peptide(vaccine,2μg/μL)was subcutaneously injected into the neck of an amyloid precursor protein/presenilin-1/tau transgenic(3×Tg-AD)mouse model.Aβ3–10-KLH peptide was re-injected at 1.5,2.5,3.5,4.5,5.5,and 6.5 months of age.Serum levels of Aβantibody were detected by enzyme-linked immunosorbent assay,while spatial learning and memory ability were evaluated by Morris water maze.Immunohistochemistry was used to detect total tau with HT7 and phosphorylated tau with AT8(phosphorylation sites Ser202 and Thr205)and AT180(phosphorylation site Thr231)antibodies in the hippocampus.In addition,western blot analysis was used to quantify AT8 and AT180 expression in the hippocampus.The results showed that after vaccine injection,mice produced high levels of Aβantibody,cognitive function was significantly improved,and total tau and phosphorylated tau levels were significantly reduced.These findings suggest that early active immunization with Aβ3–10-KLH vaccine can greatly reduce tau phosphorylation,thereby mitigating the cognitive decline of 3×Tg-AD mice.This study was approved by the Animal Ethics Committee of China Medical University,China(approval No.103-316)on April 2,2016.展开更多
Distribution and activity of mitochondda are key factors in neuronal development, synaptic plasticity and axogenesis. The majority of energy sources, necessary for cellular functions, originate from oxidative phosphor...Distribution and activity of mitochondda are key factors in neuronal development, synaptic plasticity and axogenesis. The majority of energy sources, necessary for cellular functions, originate from oxidative phosphorylation located in the inner mitochondrial membrane. The adenosine-5＇- triphosphate production is regulated by many control mechanism-firstly by oxygen, substrate level, adenosine-5＇-diphosphate level, mitochondrial membrane potential, and rate of coupling and proton leak. Recently, these mechanisms have been implemented by ＂second control mechanisms,＂ such as reversible phosphorylation of the tricarboxylic acid cycle enzymes and electron transport chain complexes, aUosteric inhibition of cytochrome c oxidase, thyroid hormones, effects of fatty acids and uncoupling proteins. Impaired function of mitochondria is implicated in many diseases ranging from mitochondrial myopathies to bipolar disorder and schizophrenia. Mitochondrial dysfunctions are usually related to the ability of mitochondria to generate adenosine-5＇-triphosphate in response to energy demands. Large amounts of reactive oxygen species are released by defective mitochondria similarly, decline of antioxidative enzyme activities （e.g. in the elderly） enhances reactive oxygen species production. We reviewed data concerning neuroplasticity, physiology, and control of mitochondrial oxidative phosphorylation and reactive oxygen species production.展开更多
Astrocytes and microglia play an orchestrated role following spinal cord injury;however,the molecular mechanisms through which microglia regulate astrocytes after spinal cord injury are not yet fully understood.Herein...Astrocytes and microglia play an orchestrated role following spinal cord injury;however,the molecular mechanisms through which microglia regulate astrocytes after spinal cord injury are not yet fully understood.Herein,microglia were pharmacologically depleted and the effects on the astrocytic response were examined.We further explored the potential mechanisms involving the signal transducers and activators of transcription 3(STAT3)pathway.For in vivo experiments,we constructed a contusion spinal cord injury model in C57BL/6 mice.To deplete microglia,all mice were treated with colony-stimulating factor 1 receptor inhibitor PLX3397,starting 2 weeks prior to surgery until they were sacrificed.Cell proliferation was examined by 5-ethynyl-2-deoxyuridine(EdU)and three pivotal inflammatory cytokines were detected by a specific Bio-Plex Pro^(TM) Reagent Kit.Locomotor function,neuroinflammation,astrocyte activation and phosphorylated STAT3(pSTAT3,a maker of activation of STAT3 signaling)levels were determined.For in vitro experiments,a microglia and astrocyte coculture system was established,and the small molecule STA21,which blocks STAT3 activation,was applied to investigate whether STAT3 signaling is involved in mediating astrocyte proliferation induced by microglia.PLX3397 administration disrupted glial scar formation,increased inflammatory spillover,induced diffuse tissue damage and impaired functional recovery after spinal cord injury.Microglial depletion markedly reduced EdU+proliferating cells,especially proliferating astrocytes at 7 days after spinal cord injury.RNA sequencing analysis showed that the JAK/STAT3 pathway was downregulated in mice treated with PLX3397.Double immunofluorescence staining confirmed that PLX3397 significantly decreased STAT3 expression in astrocytes.Importantly,in vitro coculture of astrocytes and microglia showed that microglia-induced astrocyte proliferation was abolished by STA21 administration.These findings suggest that microglial depletion impaired astrocyte proliferation and astrocytic scar formation,and induced inflammatory diffusion partly by inhibiting STAT3 phosphorylation in astrocytes following spinal cord injury.展开更多
LHCII is a crucial light-harvesting pigment/protein complex in photosystem II (PSII) supercomplex. It also participates in the light energy redistribution between photosystems and in the photoprotection via its revers...LHCII is a crucial light-harvesting pigment/protein complex in photosystem II (PSII) supercomplex. It also participates in the light energy redistribution between photosystems and in the photoprotection via its reversible dissociation with PSII and PSI (photosystem I). This reversible detachment of LHCII is regulated by phosphorylation of its own and PSII core protein. Under low light conditions, LHCII is phosphorylated and dissociated with PSII core protein complex and combined with PSI, which balances the excitation energy between PSII and PSI;Under high light environment, the phosphorylation of PSII core proteins makes LHCII detach from PSII. The dissociated LHCII presents in a free state, which involves in the thermal dissipation of excess excitation energy. During photodamage, dual phosphorylations of both PSII core proteins and LHCII complexes occur. The phosphorylation of D1 is conductive to the disintegration of photodamaged PSII and the cycle of repair. In this circumstance, the phosphorylation of LHCII is induced by reactive oxygen species (ROS) and then the phosphorylated LHCII migrates to PSI, into the repair cycle of damaged PSII. The ferredoxin (Fdr) and thioredoxin (Tdr) system may play a possible central role in the phosphorylation regulation on LHCII dissociation.展开更多
Phospholipase Cγ2 (PLCγ2) plays a pivotal role in mediation of inflammatory reaction to bacterial lipopolysaccharide (LPS) as well as serves as a key target in modulatory influence of the hormone ghrelin. Here we ex...Phospholipase Cγ2 (PLCγ2) plays a pivotal role in mediation of inflammatory reaction to bacterial lipopolysaccharide (LPS) as well as serves as a key target in modulatory influence of the hormone ghrelin. Here we explore the involvement of Rac1 and its activator, guanine nucleotide exchange factor (GEF), Dock180, in mediation of PLCγ2 activation in salivary gland acinar cells in response to P. gingivalis LPS and ghrelin. We show that stimulation of the acinar cells with the LPS leads to up-regulation in Dock and PLCγ2 activation, and is reflected in the membrane translocation of Rac1 and PLCγ2, while the effect of ghrelin is manifested by the suppression in Rac1 translocation. Further, we reveal that stimulation with the LPS leads to Dock180 phosphorylation on Tyr and Ser, while the modulatory influence of ghrelin, manifested by a drop in membrane Rac1-GTP, is asso-ciated with a distinct decrease in Dock180 phosphorylation on Ser. Moreover, we demonstrate that phosphorylation on Tyr remains under the control of Src kinase and is accompanied by Dock180 membrane translocation, while protein kinase Cδ(PKCδ) is involved in the LPS-induced phosphorylation of the membrane-recruited Dock180 on Ser. Thus, our findings underscore the role of Src/PKCδ-mediated GEF Dock180 phosphorylation on Tyr/Ser in modulation of salivary gland acinar cell PLCγ2 activation in response to P. gingivalis as well as ghrelin.展开更多
Cohesin is an evolutionary conserved complex that controls chromosome segregation during mitosis. Here we show that, in response to DNA damage, Saccharomyces cerevisiae Scc3, one of the major regulatory subunits of th...Cohesin is an evolutionary conserved complex that controls chromosome segregation during mitosis. Here we show that, in response to DNA damage, Saccharomyces cerevisiae Scc3, one of the major regulatory subunits of the Smc1-Smc3-Scc1 cohesin ring, is phosphorylated on S/T-Q residues. This event depended on the Mec1 checkpoint kinase as well as on cell cycle arrest triggered by the DNA damage checkpoint network. This phosphorylation event also took place during mitosis of an unperturbed cell cycle. The present finding that S. cerevisiae Scc3 is phosphorylated during mitosis represents a potentially important new regulatory step in cohesin’s mitotic functions.展开更多
Transient brain ischemia has been shown to induce hyperphosphorylation of the micro- tubule-associated protein tau. To further determine the mechanisms underlying these processes, we investigated the interaction betwe...Transient brain ischemia has been shown to induce hyperphosphorylation of the micro- tubule-associated protein tau. To further determine the mechanisms underlying these processes, we investigated the interaction between tau, glycogen synthase kinase （GSK）-313 and protein phos- phatase 2A. The results confirmed that tau protein was dephosphorylated during brain ischemia; in addition, the activity of GSK-3β was increased and the activity of protein phosphatase 2A was de- creased. After reperfusion, tau protein was hyperphosphorylated, the activity of GSK-3β was de- creased and the activity of protein phosphatase 2A remained low. Importantly, the interaction of tau with GSK-3β and protein phosphatase 2A was altered during ischemia and reperfusion. Lithium chloride could affect tau phosphorylation by regulating the interaction of tau with GSK-3β and pro- tein phosphatase 2A, and improve learning and memory ability of rats after transient brain ischemia. The present study demonstrated that it was the interaction of tau with GSK-3β and protein phos- phatase 2A, rather than their individual activities, that dominates the phosphorylation of tau in tran- sient brain ischemia. Hyperphosphorylated tau protein may play an important role in the evolution of brain injury in ischemic stroke. The neuroprotective effects of lithium chloride partly depend on the inhibition of tau phosphorylation during transient brain ischemia.展开更多
Objectives:Weaning induces oxidative stress in pigs,increasing the risk of diarrhea and death.Intestinal damage is associated with obstructed intestinal cell cycles.To stop damage caused by reactive oxygen species(ROS...Objectives:Weaning induces oxidative stress in pigs,increasing the risk of diarrhea and death.Intestinal damage is associated with obstructed intestinal cell cycles.To stop damage caused by reactive oxygen species(ROS),N-acetyl cysteine(NAC)has been widely employed.In this study,we examined changes in the intestinal cyclin of weaning piglets and assessed the impact of NAC on intestinal cell cycle arrest and intracellular signaling pathways.Methods:We conducted two animal experiments.In the first,we divided 12 litters of 120 newborn piglets into two groups:a control group and a weaning group.The control piglets were allowed to suckle normally.The weaning group was weaned after 3 weeks and fed a normal diet for piglets.We slaughtered six piglets from the control group and six from the weaning group.We observed cyclin changes and intestinal development at days 0,1,4,and 7 after weaning.In the second experiment,we divided 15 litters of 150 piglets that were 2 weeks old into three groups:the control group,the weaning group,and the NAC group.Control piglets were allowed to suckle normally.Piglets in the weaning and NAC groups were weaned when they were 21 days old.The NAC group was fed a basal diet supplemented with 500 mg/kg NAC,and the weaning group was fed the basal diet alone.The experimental period was 14–25 days of age.Four days after weaning,we slaughtered one piglet from each litter.We then analyzed intestinal cell cycle indexes,intestinal oxidative stress,c-Jun N-terminal kinase(JNK),extracellular signal-regulated kinase(ERK),and p38 phosphorylation.Results:Weaning decreased the piglets’feed intake and daily gain,reduced the serum antioxidant capacity,and increased the intestinal ROS level.Furthermore,the jejunum histology and barrier development of the jejunum exhibited damage after weaning,the microvilli displayed hypoplasia,and the p21 and p27 protein expression levels of the jejunum were significantly elevated.We did not observe any significant differences in cyclin D and E after days 1,4,and 7 post-weaning compared with the control group.We observed,however,significantly increased cyclin D and E expression,lower ERK,JNK,and p38 kinase phosphorylation;villus atrophy alleviation;decreased p21 and p27 expression;and increased average daily intake of feed and weight gain.Conclusion:This research demonstrates that weaning stress inhibits piglet intestinal proliferation by reducing cyclin D and cyclin E expression.NAC downregulates p21 and p27 through modulating mitogen-activated protein kinases(MAPKase)phosphorylation,thereby promoting cell proliferation.The results indicate that NAC promotes intestinal function and the integrity of enterocytes and holds promise as a new feed additive for animal health.展开更多
Sequence similarities were found between protein and DNA sequences encoding certain part of conserved variable immunoglobulin domains (i.e. conserved IgV) and phosphorylation sites. Hypermutation motifs were then indi...Sequence similarities were found between protein and DNA sequences encoding certain part of conserved variable immunoglobulin domains (i.e. conserved IgV) and phosphorylation sites. Hypermutation motifs were then indicated in the majority of the corresponding non-IgV nucleotide sequences. According to database confirmations or double prediction of phosphorylation sites, 80% of the selected human and mouse IgV-related phosphorylation sites or their highly probable candidates exhibited substrate relationship to ataxia-telangiectasia-mutated kinase known as ATM. In accordance with literature data, inactivation of ATM by mutations can participate in the mechanisms of carcinogenesis, neurodegeneration and possibly also in aging. In agreement with this relationship, some of the selected IgV-/ATM-related segments formed molecules specifically involved in carcinogenesis. The selected IgV-related sequence segments were also similar to certain segments of higher plants containing immunoglobulin-like repeats and related regions. Bioinformatic analysis of some selected plant sequences then indicated the presence of catalytic domains composing serine/threonine/tyrosine receptor/receptor-like kinases, which are considered important structures for evolution of very early and part of later Ig-domain-related immunity. The analyzed conserved domain similarities also suggested certain interesting structural and phylogenic relationships, which need to be further investigated. This review in fact briefly summarizes the findings on the subject from the last twenty years.展开更多
Alzheimer’s disease is a neurodegenerative disorder characterized by the amyloid accumulation in the brains of patients with Alzheimer’s disease.The pathogenesis of Alzheimer’s disease is mainly mediated by the pho...Alzheimer’s disease is a neurodegenerative disorder characterized by the amyloid accumulation in the brains of patients with Alzheimer’s disease.The pathogenesis of Alzheimer’s disease is mainly mediated by the phosphorylation and aggregation of tau protein.Among the multiple causes of tau hyperphosphorylation,brain insulin resistance has generated much attention,and inositols as insulin sensitizers,are currently considered candidates for drug development.The present narrative review revises the interactions between these three elements:Alzheimer’s disease-tau-inositols,which can eventually identify targets for new disease modifiers capable of bringing hope to the millions of people affected by this devastating disease.展开更多
Objective:To observe the effect of Anshen Dingzhi Decoction on tau phosphorylation in hippocampus of Alzheimer's disease rat model and its related mechanism.Methods:SD rats were randomly divided into control group...Objective:To observe the effect of Anshen Dingzhi Decoction on tau phosphorylation in hippocampus of Alzheimer's disease rat model and its related mechanism.Methods:SD rats were randomly divided into control group,model group,Anshen Dingzhi Decoction(low,medium and high dose group)and Dopazide group.Except for the control group,the rats in the other groups were injected with streptozotocin into the lateral ventricle to replicate the model of Alzheimer's disease and then given corresponding drugs for 2 weeks.orris water maze test was used to detect learning and memory ability of rats,HE staining was used to detect hippocampal histological morphology,Western-blot was used to detect tau phosphorylation level and expression abundance of BDNF and TrkB protein in hippocampal tissue of rats.Results:The escape latency of the model group was significantly increased,the number of crossing platforms and effective residence time were significantly reduced,the phosphorylation level of tau protein was significantly increased,and the phosphorylation levels of BDNF and TrkB protein were significantly decreased when compared with the control group,the difference has statistically significant(P<0.05).Anshen Dingzhi Fang could significantly reduce the escape latency of AD rats,increase the number of crossing platforms and effective residence time,inhibit the phosphorylation of tau protein,and up-regulate the phosphorylation of BDNF and TrkB protein,the difference was statistically significant when compared with the model group(P<0.05).Conclusion:Anshen Dingzhi Fang can inhibit the phosphorylation of tau protein by activating BDNF/TrkB signaling pathway and promote the learning and memory ability of AD rats.展开更多
Meiotic resumption in mammalian oocytes involves nuclear and organelle structural changes,notably the chromatin configuration transition from a non-surrounding nucleolus(NSN)to surrounding nucleolus(SN)in germinal ves...Meiotic resumption in mammalian oocytes involves nuclear and organelle structural changes,notably the chromatin configuration transition from a non-surrounding nucleolus(NSN)to surrounding nucleolus(SN)in germinal vesicle oocytes.In the current study,we found that nuclear speckles(NSs),a subnuclear structure mainly composed of serine-arginine(SR)proteins,changed from a diffuse spotted distribution in mouse NSN oocytes to an aggregated pattern in SN oocytes.We also found that the SR protein-specific kinase 1(SRPK1),an enzyme that phosphorylates SR proteins,co-localized with NSs at the SN stage,and that NSN oocytes failed to transition to SN oocytes after the inhibition of SRPK1 activity.Furthermore,the typical structure of the chromatin ring around the nucleolus in SN oocytes collapsed after treatment with an SRPK1 inhibitor.Mechanistically,phosphorylated SR proteins were found to be related to chromatin as shown by a salt extraction experiment,and in situ DNaseⅠassay showed that the accessibility of chromatin was enhanced in SN oocytes when SRPK1 was inhibited,accompanied by a decreased repressive modification on histone and the abnormal recurrence of a transcriptional signal.In conclusion,our results indicated that SRPK1-regulated phosphorylation of SR proteins was involved in the NSN-SN transition and played an important role in maintaining the condensed nucleus of SN oocytes via interacting with chromatin.展开更多
Nowadays,the cumulative intake of glucocorticoids has become the most common pathogenic factor for non-traumatic osteonecrosis of the femoral head(ONFH).Apoptosis of osteoblasts is considered as the main reason of ONF...Nowadays,the cumulative intake of glucocorticoids has become the most common pathogenic factor for non-traumatic osteonecrosis of the femoral head(ONFH).Apoptosis of osteoblasts is considered as the main reason of ONFH at the molecular level.Glycogen synthase kinase 3β(GSK3β)is an important regulator of cellular differentiation and apoptosis pathway,which can modulate the balance between osteoblasts and osteoclasts.Several studies have reported about its function in osteoporosis,but little is known about it in osteonecrosis.In our study,lipopolysaccharide and methylprednisolone were utilized to establish a rat ONFH model.The phosphorylation of GSK3βSer-9 was decreased in the model.Western blotting examination ofβ-catenin,Bcl-2,Bax and caspase-3 revealed that the osteoblasts were apoptotic.In dexamethasone(Dex)-incubated primary osteoblasts,the expression profile of GSK3βphosphorylation and apoptotic factors were consistent with those in the rat ONFH model.To further investigate the regulation of osteonecrosis caused by GSK3β,the expression and function of GSK3βwere inhibited in Dex-incubated primary osteoblasts.The knockdown of GSK3βby siRNA decreased the expression of Bax and cleaved caspase-3,but increased Bcl-2 andβ-catenin.On the other hand,selective inhibition of GSK3βfunction by LiCl counteracted the activation of caspase-3 induced by Dex.Our work is the first study about the GSK3P phosphorylation in ONFH,and provides evidence for further therapeutic methods.展开更多
Phosphorylation of protein is an important post-translational modification that enables activation of various enzymes and receptors included in signaling pathways. To reduce the cost of identifying phosphorylation sit...Phosphorylation of protein is an important post-translational modification that enables activation of various enzymes and receptors included in signaling pathways. To reduce the cost of identifying phosphorylation site by laborious experiments, computational prediction of it has been actively studied. In this study, by adopting a new set of features and applying feature selection by Random Forest with grid search before training by Support Vector Machine, our method achieved better or comparable performance of phosphorylation site prediction for two different data sets.展开更多
BACKGROUND Intestinal inflammation and epithelial injury are the leading actors of inflammatory bowel disease(IBD),causing an excessive pro-inflammatory cytokines expression.Tristetraprolin(TTP),an mRNA binding protei...BACKGROUND Intestinal inflammation and epithelial injury are the leading actors of inflammatory bowel disease(IBD),causing an excessive pro-inflammatory cytokines expression.Tristetraprolin(TTP),an mRNA binding protein,plays a role in regulating the inflammatory factors,recognizing specific sequences on the 3’untranslated region of cytokine mRNAs.TTP activity depends on its phosphorylation state:the unphosphorylated TTP degrades pro-inflammatory cytokine mRNAs;on the contrary,the phosphorylated TTP fails to destabilize mRNAs furthering their expression.The phospho-TTP forms a complex with the chaperone protein 14-3-3.This binding could be one of the factors that promote intestinal inflammation as a cause of disease progression.AIM To assess if TTP phosphorylation has a role in paediatric IBD.METHODS The study was carried out on a cohort of paediatric IBD patients.For each patient enrolled,a specimen of inflamed and non-inflamed colonic mucosa was collected.Furthermore,the experiments were conducted on macrophages differentiated from blood samples of the same patients.Macrophages from healthy donors’blood were used as controls.Co-immunoprecipitation assay and immunoblotting analyses were performed to observe the formation of the phospho-TTP/14-3-3 complex.In the same samples TNF-αexpression was also evaluated as major factor of the pro-inflammatory activity.RESULTS In this work we studied indirectly the phosphorylation of TTP through the binding with the chaperone protein 14-3-3.In inflamed and non-inflamed colon mucosa of IBD paediatric patients immunoblot assay demonstrated a higher expression of the TTP in inflamed samples respect to the non-inflamed;the coimmunoprecipitated 14-3-3 protein showed the same trend of expression.In the TNF-αgene expression analysis higher levels of the cytokine in inflamed tissues compared to controls were evident.The same experiments were conducted on macrophages from IBD paediatric patients and healthy controls.The immunoblot results demonstrated a high expression of both TTP and co-immunoprecipitated 14-4-3 protein in IBD-derived macrophages in comparison to healthy donors.TNF-αprotein levels from macrophages lysates showed the same trend of expression in favour of IBD paediatric patients compared to healthy controls.CONCLUSION In this work,for the first time,we describe a relation between phospho-TTP/14-3-3 complex and IBD.Indeed,a higher expression of TTP/14-3-3 was recorded in IBD samples in comparison to controls.展开更多
Disturbances in constitutive nitric oxide synthase (cNOS) activation associated with H. pylori colonization of gastric mucosa are considered of major consequences in defining the extent of inflammatory involvement. As...Disturbances in constitutive nitric oxide synthase (cNOS) activation associated with H. pylori colonization of gastric mucosa are considered of major consequences in defining the extent of inflammatory involvement. As rapid changes in cNOS activation are linked to the enzyme phosphorylation at the specific Ser/Thr residues, we investigated the influence of H. pylori LPS and gastric hormone, ghrelin, on the processes of phosphorylation of these two critical sites in gastric mucosal cells. We show that the LPS-induced reduction in cNOS activity is reflected in the phosphorylation on Thr497, while the countering effect of ghrelin is associated with a rapid increase in cNOS phosphorylation on Ser1179. Further, we demonstrate that cNOS phosphorylation on Thr497 as well as Ser1179 displays dependence on PKCδ. However, while the LPS-induced suppression in cNOS activation shows reliance on the phosphorylation of PKCδ and PI3K on Ser, the effect of ghrelin is manifested by the increase in phosphorylation of PKCδ and PI3K on Tyr, as well as membrane translocation and phosphorylation of Akt on Ser493. Thus, our findings suggest that the LPS-induced suppression in cNOS activation is mediated by PKCδ-controlled phosphorylation of PI3K on Ser that interferes with the membrane recruitment of Akt and promotes cNOS phosphorylation on Thr497, and that ghrelin-elicited up-regulation in cNOS activation relies on the PKCδ-directed phosphorylation of PI3K on Tyr that stimulates the membrane localization of Akt and enhances cNOS phosphorylation on Ser1179.展开更多
The signaling events underlying oral mucosal inflammatory responses to P. gingivalis and its key endotoxin, lipopolysaccharide (LPS), relay primarily on the LPS engagement of Toll-like receptor-4 (TLR4), and the activ...The signaling events underlying oral mucosal inflammatory responses to P. gingivalis and its key endotoxin, lipopolysaccharide (LPS), relay primarily on the LPS engagement of Toll-like receptor-4 (TLR4), and the activation of IκB-kinase complex (IKK) and mitogen-activated protein kinases (MAPKs that exert their control over transcription factors implicated in the regulation of iNOS and COX-2 proinflammatory genes expression). Since spleen tyrosine kinase (Syk) has emerged recently as a major amplifier in the production of proinflammatory mediators, we investigated the process of recruitment and interaction of Syk with TLR4 in salivary gland acinar cells in response to P. gingivalis LPS. Our findings revealed that stimulation of the acinar cells with the LPS leads to protein kinase Cδ (PKCδ)-mediated phosphorylation of Syk on Ser which results in its localization with the membrane associated TLR4 complex and the activation through phosphorylation on Tyr. Further, our results support the involvement of Syk in the amplification of transcription factors involved in the assembly and expression of transcription complexes associated with the induction in COX-2 and iNOS genes. Therefore, our data suggest that PKCδ is a primary linchpin affecting the Syk recruitment to the membrane localized TLR4, and hence affects the efficiency of the kinase activation and the magnitude of oral mucosal inflammatory response to P. gingivalis.展开更多
文摘AIM： To study the relationship between interleukin-lbeta （IL-1β） up-regulating tissue inhibitor of matrix metalloproteinase-1 （TIMMP-1） mRNA expression and phosphorylation of both c-jun N-terminal kinase （INK） and p38 in rat heffatic stellate cells （HSC）. METHODS： RT-PCR was performed to measure the expression of TIMMP-1 mRNA in rat HSC. Western blot was performed to measure IL-1β-induced JNK and p38 activities in rat HSC. RESULTS： TIMMP-1 mRNA expression （1.191± 0.079） was much higher after treatment with IL-1β （10 ng/mL） for 24 h than in control group （0.545±0.091） （P〈0.01）. IL-1β activated INK and p38 in a time-dependent manner. After stimulation with IL-1β for 0, 5, 15, 30, 60 and 120 min, the INK activity was 0.982±0.299, 1.501±0.720, 2.133±0.882, 3.360±0.452, 2.181±0.789, and 1.385 ± 0.368, respectively. There was a significant difference in JNK activity at 15 min （P〈 0.01）, 30 min （P〈 0.01） and 60 min （P〈0.01） in comparison to that at 0 min. The p38 activity was 1.061±0.310, 2.050±0.863, 2.380±0.573, 2.973±0.953, 2.421±0.793, and 1.755 ± 0.433 at the 6 time points （0, 5, 15, 30, 60 and 120 min） respectively. There was a significant difference in p38 activity at 5 min （P〈0.05）, 15 min （P〈0.01）, 30 min （P〈0.01） and 60 min （P〈0.01） compared to that at 0 min. TIMMP-1 mRNA expression trended to decrease in 3 groups pretreated with different concentrations of SP600125 （10 μmol/L, 1.022±0.113; 20 μmol/L, 0.869±0.070; 40 μmol/L, 0.666±0.123）. Their decreases were all significant （P〈0.05, P〈0.01, P〈0.01） in comparison to control group （without SP600125 treatment, 1.163±0.107）. In the other 3 groups pretreated with different concentrations of SB203580 （10 μmol/L, 1.507±0.099; 20 μmol/L, 1.698±0.107; 40 μmol/L, 1.857±0.054）, the expression of TIMMP-1 mRNA increased. Their levels were higher than those in the control group （without SB203580 treatment, 1.027 ± 0.061） with a significant statistical significance （P〈 0.01）. CONCLUSION： IL-1β has a direct action on hepatic fibrosis by up-regulating TIMMP-1 mRNA expression in ratessionin in rate HSC.JNK and p38 mitogen-activated protein kinases （MAPKs） are involved in IL-1β-induced TIMMP-1 gene expression, and play a distinct role in this process, indicating that p38 and .INK pathways cooperatively mediate TIMP-1 mRNA expression in rat HSC.
文摘Our previous studies showed a predominance of high molecular weight protein group in tumor nuclear matrices. Contrary to normal cells, proteins of this group are preferentially phosphorylated. Phosphoproteins of hepatoma nuclear matrix are selectively subjected to rapid proteolysis. By alkali treatment and a monoclonal antibody against phosphotyrosyl residue the presence of two high molecular weight bands of phosphotyrosyl-containing proteins was detected in nuclear matrices of tumor but not of normal liver cells. High molecular weight protein group of tumor nuclear matrices revealed also a rapid turnover and preferential incorporation of labeled amino acids selectively inhibited by chloramphenicol.
基金the National Natural Science Foundation of China,No.81679154,No.81871547.
文摘BACKGROUND Intestinal ischemia reperfusion(I/R)occurs in various diseases,such as trauma and intestinal transplantation.Excessive reactive oxygen species(ROS)accumulation and subsequent apoptotic cell death in intestinal epithelia are important causes of I/R injury.PTEN-induced putative kinase 1(PINK1)and phosphorylation of dynamin-related protein 1(DRP1)are critical regulators of ROS and apoptosis.However,the correlation of PINK1 and DRP1 and their function in intestinal I/R injury have not been investigated.Thus,examining the PINK1/DRP1 pathway may help to identify a protective strategy and improve the patient prognosis.AIM To clarify the mechanism of the PINK1/DRP1 pathway in intestinal I/R injury.METHODS Male C57BL/6 mice were used to generate an intestinal I/R model via superior mesenteric artery occlusion followed by reperfusion.Chiu’s score was used to evaluate intestinal mucosa damage.The mitochondrial fission inhibitor mdivi-1 was administered by intraperitoneal injection.Caco-2 cells were incubated in vitro in hypoxia/reoxygenation conditions.Small interfering RNAs and overexpression plasmids were transfected to regulate PINK1 expression.The protein expression levels of PINK1,DRP1,p-DRP1 and cleaved caspase 3 were measured by Western blotting.Cell viability was evaluated using a Cell Counting Kit-8 assay and cell apoptosis was analyzed by TUNEL staining.Mitochondrial fission and ROS were tested by MitoTracker and MitoSOX respectively.RESULTS Intestinal I/R and Caco-2 cell hypoxia/reoxygenation decreased the expression of PINK1 and p-DRP1 Ser637.Pretreatment with mdivi-1 inhibited mitochondrial fission,ROS generation,and apoptosis and ameliorated cell injury in intestinal I/R.Upon PINK1 knockdown or overexpression in vitro,we found that p-DRP1 Ser637 expression and DRP1 recruitment to the mitochondria were associated with PINK1.Furthermore,we verified the physical combination of PINK1 and p-DRP1 Ser637.CONCLUSION PINK1 is correlated with mitochondrial fission and apoptosis by regulating DRP1 phosphorylation in intestinal I/R.These results suggest that the PINK1/DRP1 pathway is involved in intestinal I/R injury,and provide a new approach for prevention and treatment.
基金supported by the National Natural Science Foundation of China,No.81371227(to YPC)
文摘Active and passive anti-Aβimmunotherapies have successfully been used for the prevention and treatment of Alzheimer’s disease animal models.However,clinical use of these immunotherapies is not effective,because the vaccination is administered too late.At 1 month of age,100μL of Aβ3–10-KLH peptide(vaccine,2μg/μL)was subcutaneously injected into the neck of an amyloid precursor protein/presenilin-1/tau transgenic(3×Tg-AD)mouse model.Aβ3–10-KLH peptide was re-injected at 1.5,2.5,3.5,4.5,5.5,and 6.5 months of age.Serum levels of Aβantibody were detected by enzyme-linked immunosorbent assay,while spatial learning and memory ability were evaluated by Morris water maze.Immunohistochemistry was used to detect total tau with HT7 and phosphorylated tau with AT8(phosphorylation sites Ser202 and Thr205)and AT180(phosphorylation site Thr231)antibodies in the hippocampus.In addition,western blot analysis was used to quantify AT8 and AT180 expression in the hippocampus.The results showed that after vaccine injection,mice produced high levels of Aβantibody,cognitive function was significantly improved,and total tau and phosphorylated tau levels were significantly reduced.These findings suggest that early active immunization with Aβ3–10-KLH vaccine can greatly reduce tau phosphorylation,thereby mitigating the cognitive decline of 3×Tg-AD mice.This study was approved by the Animal Ethics Committee of China Medical University,China(approval No.103-316)on April 2,2016.
基金supported by grant NoMSM0021620849 given by the Ministry of Education,Youth and Sports of the Czech Republicby project PRVOUK-P26/LF1/4given by Charles University in Prague+1 种基金by grant No. SVV-2012-264514 from Charles University in Pragueby grant No.41310 given by the Grant Agency of Charles University in Prague,Czech Republic
文摘Distribution and activity of mitochondda are key factors in neuronal development, synaptic plasticity and axogenesis. The majority of energy sources, necessary for cellular functions, originate from oxidative phosphorylation located in the inner mitochondrial membrane. The adenosine-5＇- triphosphate production is regulated by many control mechanism-firstly by oxygen, substrate level, adenosine-5＇-diphosphate level, mitochondrial membrane potential, and rate of coupling and proton leak. Recently, these mechanisms have been implemented by ＂second control mechanisms,＂ such as reversible phosphorylation of the tricarboxylic acid cycle enzymes and electron transport chain complexes, aUosteric inhibition of cytochrome c oxidase, thyroid hormones, effects of fatty acids and uncoupling proteins. Impaired function of mitochondria is implicated in many diseases ranging from mitochondrial myopathies to bipolar disorder and schizophrenia. Mitochondrial dysfunctions are usually related to the ability of mitochondria to generate adenosine-5＇-triphosphate in response to energy demands. Large amounts of reactive oxygen species are released by defective mitochondria similarly, decline of antioxidative enzyme activities （e.g. in the elderly） enhances reactive oxygen species production. We reviewed data concerning neuroplasticity, physiology, and control of mitochondrial oxidative phosphorylation and reactive oxygen species production.
基金supported by the Natural Science Foundation of Guangdong Province,No.2020A1515010090(to ZLZ)the Science and Technology Project Foundation of Guangzhou City,No.202002030004(to HZ).
文摘Astrocytes and microglia play an orchestrated role following spinal cord injury;however,the molecular mechanisms through which microglia regulate astrocytes after spinal cord injury are not yet fully understood.Herein,microglia were pharmacologically depleted and the effects on the astrocytic response were examined.We further explored the potential mechanisms involving the signal transducers and activators of transcription 3(STAT3)pathway.For in vivo experiments,we constructed a contusion spinal cord injury model in C57BL/6 mice.To deplete microglia,all mice were treated with colony-stimulating factor 1 receptor inhibitor PLX3397,starting 2 weeks prior to surgery until they were sacrificed.Cell proliferation was examined by 5-ethynyl-2-deoxyuridine(EdU)and three pivotal inflammatory cytokines were detected by a specific Bio-Plex Pro^(TM) Reagent Kit.Locomotor function,neuroinflammation,astrocyte activation and phosphorylated STAT3(pSTAT3,a maker of activation of STAT3 signaling)levels were determined.For in vitro experiments,a microglia and astrocyte coculture system was established,and the small molecule STA21,which blocks STAT3 activation,was applied to investigate whether STAT3 signaling is involved in mediating astrocyte proliferation induced by microglia.PLX3397 administration disrupted glial scar formation,increased inflammatory spillover,induced diffuse tissue damage and impaired functional recovery after spinal cord injury.Microglial depletion markedly reduced EdU+proliferating cells,especially proliferating astrocytes at 7 days after spinal cord injury.RNA sequencing analysis showed that the JAK/STAT3 pathway was downregulated in mice treated with PLX3397.Double immunofluorescence staining confirmed that PLX3397 significantly decreased STAT3 expression in astrocytes.Importantly,in vitro coculture of astrocytes and microglia showed that microglia-induced astrocyte proliferation was abolished by STA21 administration.These findings suggest that microglial depletion impaired astrocyte proliferation and astrocytic scar formation,and induced inflammatory diffusion partly by inhibiting STAT3 phosphorylation in astrocytes following spinal cord injury.
文摘LHCII is a crucial light-harvesting pigment/protein complex in photosystem II (PSII) supercomplex. It also participates in the light energy redistribution between photosystems and in the photoprotection via its reversible dissociation with PSII and PSI (photosystem I). This reversible detachment of LHCII is regulated by phosphorylation of its own and PSII core protein. Under low light conditions, LHCII is phosphorylated and dissociated with PSII core protein complex and combined with PSI, which balances the excitation energy between PSII and PSI;Under high light environment, the phosphorylation of PSII core proteins makes LHCII detach from PSII. The dissociated LHCII presents in a free state, which involves in the thermal dissipation of excess excitation energy. During photodamage, dual phosphorylations of both PSII core proteins and LHCII complexes occur. The phosphorylation of D1 is conductive to the disintegration of photodamaged PSII and the cycle of repair. In this circumstance, the phosphorylation of LHCII is induced by reactive oxygen species (ROS) and then the phosphorylated LHCII migrates to PSI, into the repair cycle of damaged PSII. The ferredoxin (Fdr) and thioredoxin (Tdr) system may play a possible central role in the phosphorylation regulation on LHCII dissociation.
文摘Phospholipase Cγ2 (PLCγ2) plays a pivotal role in mediation of inflammatory reaction to bacterial lipopolysaccharide (LPS) as well as serves as a key target in modulatory influence of the hormone ghrelin. Here we explore the involvement of Rac1 and its activator, guanine nucleotide exchange factor (GEF), Dock180, in mediation of PLCγ2 activation in salivary gland acinar cells in response to P. gingivalis LPS and ghrelin. We show that stimulation of the acinar cells with the LPS leads to up-regulation in Dock and PLCγ2 activation, and is reflected in the membrane translocation of Rac1 and PLCγ2, while the effect of ghrelin is manifested by the suppression in Rac1 translocation. Further, we reveal that stimulation with the LPS leads to Dock180 phosphorylation on Tyr and Ser, while the modulatory influence of ghrelin, manifested by a drop in membrane Rac1-GTP, is asso-ciated with a distinct decrease in Dock180 phosphorylation on Ser. Moreover, we demonstrate that phosphorylation on Tyr remains under the control of Src kinase and is accompanied by Dock180 membrane translocation, while protein kinase Cδ(PKCδ) is involved in the LPS-induced phosphorylation of the membrane-recruited Dock180 on Ser. Thus, our findings underscore the role of Src/PKCδ-mediated GEF Dock180 phosphorylation on Tyr/Ser in modulation of salivary gland acinar cell PLCγ2 activation in response to P. gingivalis as well as ghrelin.
文摘Cohesin is an evolutionary conserved complex that controls chromosome segregation during mitosis. Here we show that, in response to DNA damage, Saccharomyces cerevisiae Scc3, one of the major regulatory subunits of the Smc1-Smc3-Scc1 cohesin ring, is phosphorylated on S/T-Q residues. This event depended on the Mec1 checkpoint kinase as well as on cell cycle arrest triggered by the DNA damage checkpoint network. This phosphorylation event also took place during mitosis of an unperturbed cell cycle. The present finding that S. cerevisiae Scc3 is phosphorylated during mitosis represents a potentially important new regulatory step in cohesin’s mitotic functions.
基金supported by the National High Technology Research and Development Program of China(863 Program),No.2012AA020905the Biological Industry Development Funds of Shenzhen,No.JC201005260093A+1 种基金the National Natural Science Foundation of China/Research Grants Council Joint Research Scheme,No.81161160570the National Natural Science Foundation of China,No.81171143the Tsinghua-Yue-Yuen Medical Sciences Fund
文摘Transient brain ischemia has been shown to induce hyperphosphorylation of the micro- tubule-associated protein tau. To further determine the mechanisms underlying these processes, we investigated the interaction between tau, glycogen synthase kinase （GSK）-313 and protein phos- phatase 2A. The results confirmed that tau protein was dephosphorylated during brain ischemia; in addition, the activity of GSK-3β was increased and the activity of protein phosphatase 2A was de- creased. After reperfusion, tau protein was hyperphosphorylated, the activity of GSK-3β was de- creased and the activity of protein phosphatase 2A remained low. Importantly, the interaction of tau with GSK-3β and protein phosphatase 2A was altered during ischemia and reperfusion. Lithium chloride could affect tau phosphorylation by regulating the interaction of tau with GSK-3β and pro- tein phosphatase 2A, and improve learning and memory ability of rats after transient brain ischemia. The present study demonstrated that it was the interaction of tau with GSK-3β and protein phos- phatase 2A, rather than their individual activities, that dominates the phosphorylation of tau in tran- sient brain ischemia. Hyperphosphorylated tau protein may play an important role in the evolution of brain injury in ischemic stroke. The neuroprotective effects of lithium chloride partly depend on the inhibition of tau phosphorylation during transient brain ischemia.
基金supported by the Jilin Agricultural Science and Technology University under the Scientific Startup Foundation for Doctors((2022)733)Shanghai Jiao Tong University under the National Natural Science Foundation of China(30972103).
文摘Objectives:Weaning induces oxidative stress in pigs,increasing the risk of diarrhea and death.Intestinal damage is associated with obstructed intestinal cell cycles.To stop damage caused by reactive oxygen species(ROS),N-acetyl cysteine(NAC)has been widely employed.In this study,we examined changes in the intestinal cyclin of weaning piglets and assessed the impact of NAC on intestinal cell cycle arrest and intracellular signaling pathways.Methods:We conducted two animal experiments.In the first,we divided 12 litters of 120 newborn piglets into two groups:a control group and a weaning group.The control piglets were allowed to suckle normally.The weaning group was weaned after 3 weeks and fed a normal diet for piglets.We slaughtered six piglets from the control group and six from the weaning group.We observed cyclin changes and intestinal development at days 0,1,4,and 7 after weaning.In the second experiment,we divided 15 litters of 150 piglets that were 2 weeks old into three groups:the control group,the weaning group,and the NAC group.Control piglets were allowed to suckle normally.Piglets in the weaning and NAC groups were weaned when they were 21 days old.The NAC group was fed a basal diet supplemented with 500 mg/kg NAC,and the weaning group was fed the basal diet alone.The experimental period was 14–25 days of age.Four days after weaning,we slaughtered one piglet from each litter.We then analyzed intestinal cell cycle indexes,intestinal oxidative stress,c-Jun N-terminal kinase(JNK),extracellular signal-regulated kinase(ERK),and p38 phosphorylation.Results:Weaning decreased the piglets’feed intake and daily gain,reduced the serum antioxidant capacity,and increased the intestinal ROS level.Furthermore,the jejunum histology and barrier development of the jejunum exhibited damage after weaning,the microvilli displayed hypoplasia,and the p21 and p27 protein expression levels of the jejunum were significantly elevated.We did not observe any significant differences in cyclin D and E after days 1,4,and 7 post-weaning compared with the control group.We observed,however,significantly increased cyclin D and E expression,lower ERK,JNK,and p38 kinase phosphorylation;villus atrophy alleviation;decreased p21 and p27 expression;and increased average daily intake of feed and weight gain.Conclusion:This research demonstrates that weaning stress inhibits piglet intestinal proliferation by reducing cyclin D and cyclin E expression.NAC downregulates p21 and p27 through modulating mitogen-activated protein kinases(MAPKase)phosphorylation,thereby promoting cell proliferation.The results indicate that NAC promotes intestinal function and the integrity of enterocytes and holds promise as a new feed additive for animal health.
文摘Sequence similarities were found between protein and DNA sequences encoding certain part of conserved variable immunoglobulin domains (i.e. conserved IgV) and phosphorylation sites. Hypermutation motifs were then indicated in the majority of the corresponding non-IgV nucleotide sequences. According to database confirmations or double prediction of phosphorylation sites, 80% of the selected human and mouse IgV-related phosphorylation sites or their highly probable candidates exhibited substrate relationship to ataxia-telangiectasia-mutated kinase known as ATM. In accordance with literature data, inactivation of ATM by mutations can participate in the mechanisms of carcinogenesis, neurodegeneration and possibly also in aging. In agreement with this relationship, some of the selected IgV-/ATM-related segments formed molecules specifically involved in carcinogenesis. The selected IgV-related sequence segments were also similar to certain segments of higher plants containing immunoglobulin-like repeats and related regions. Bioinformatic analysis of some selected plant sequences then indicated the presence of catalytic domains composing serine/threonine/tyrosine receptor/receptor-like kinases, which are considered important structures for evolution of very early and part of later Ig-domain-related immunity. The analyzed conserved domain similarities also suggested certain interesting structural and phylogenic relationships, which need to be further investigated. This review in fact briefly summarizes the findings on the subject from the last twenty years.
基金supported by the European Regional Development Funds-European Union(ERDF-EU),FATZHEIMER project(EU-LAC HEALTH 2020,16/T010131 to FRdF),“Una manera de hacer Europa”Ministerio de Economía,Industria y Competitividad,Gobierno de Espa?a,Programa Estatal de Investigación,Desarrollo e Innovación Orientada a los Retos de la Sociedad(RTC2019-007329-1 to FRdF)+2 种基金Consejería de Economía,Conocimiento y Universidad,Junta de Andalucía,Plan Andaluz de Investigación,Desarrollo e Innovación(P18TP-5194 to FRdF)Instituto de Salud CarlosⅢ(DTS22/00021 to FRdF)DMV(FI20/00227)holds a“PFIS’’predoctoral contract from the National System of Health,EU-ERDF-Instituto de Salud CarlosⅢ。
文摘Alzheimer’s disease is a neurodegenerative disorder characterized by the amyloid accumulation in the brains of patients with Alzheimer’s disease.The pathogenesis of Alzheimer’s disease is mainly mediated by the phosphorylation and aggregation of tau protein.Among the multiple causes of tau hyperphosphorylation,brain insulin resistance has generated much attention,and inositols as insulin sensitizers,are currently considered candidates for drug development.The present narrative review revises the interactions between these three elements:Alzheimer’s disease-tau-inositols,which can eventually identify targets for new disease modifiers capable of bringing hope to the millions of people affected by this devastating disease.
基金Natural science fund of Heilongjiang province(No.H2018063).
文摘Objective:To observe the effect of Anshen Dingzhi Decoction on tau phosphorylation in hippocampus of Alzheimer's disease rat model and its related mechanism.Methods:SD rats were randomly divided into control group,model group,Anshen Dingzhi Decoction(low,medium and high dose group)and Dopazide group.Except for the control group,the rats in the other groups were injected with streptozotocin into the lateral ventricle to replicate the model of Alzheimer's disease and then given corresponding drugs for 2 weeks.orris water maze test was used to detect learning and memory ability of rats,HE staining was used to detect hippocampal histological morphology,Western-blot was used to detect tau phosphorylation level and expression abundance of BDNF and TrkB protein in hippocampal tissue of rats.Results:The escape latency of the model group was significantly increased,the number of crossing platforms and effective residence time were significantly reduced,the phosphorylation level of tau protein was significantly increased,and the phosphorylation levels of BDNF and TrkB protein were significantly decreased when compared with the control group,the difference has statistically significant(P<0.05).Anshen Dingzhi Fang could significantly reduce the escape latency of AD rats,increase the number of crossing platforms and effective residence time,inhibit the phosphorylation of tau protein,and up-regulate the phosphorylation of BDNF and TrkB protein,the difference was statistically significant when compared with the model group(P<0.05).Conclusion:Anshen Dingzhi Fang can inhibit the phosphorylation of tau protein by activating BDNF/TrkB signaling pathway and promote the learning and memory ability of AD rats.
基金National Natural Science Foundation of China(Grant Nos.32070838 and 82301874)Open Fund of State Key Laboratory of Reproductive Medicine,Nanjing Medical University(Grant No.SKLRM K202102)。
文摘Meiotic resumption in mammalian oocytes involves nuclear and organelle structural changes,notably the chromatin configuration transition from a non-surrounding nucleolus(NSN)to surrounding nucleolus(SN)in germinal vesicle oocytes.In the current study,we found that nuclear speckles(NSs),a subnuclear structure mainly composed of serine-arginine(SR)proteins,changed from a diffuse spotted distribution in mouse NSN oocytes to an aggregated pattern in SN oocytes.We also found that the SR protein-specific kinase 1(SRPK1),an enzyme that phosphorylates SR proteins,co-localized with NSs at the SN stage,and that NSN oocytes failed to transition to SN oocytes after the inhibition of SRPK1 activity.Furthermore,the typical structure of the chromatin ring around the nucleolus in SN oocytes collapsed after treatment with an SRPK1 inhibitor.Mechanistically,phosphorylated SR proteins were found to be related to chromatin as shown by a salt extraction experiment,and in situ DNaseⅠassay showed that the accessibility of chromatin was enhanced in SN oocytes when SRPK1 was inhibited,accompanied by a decreased repressive modification on histone and the abnormal recurrence of a transcriptional signal.In conclusion,our results indicated that SRPK1-regulated phosphorylation of SR proteins was involved in the NSN-SN transition and played an important role in maintaining the condensed nucleus of SN oocytes via interacting with chromatin.
文摘Nowadays,the cumulative intake of glucocorticoids has become the most common pathogenic factor for non-traumatic osteonecrosis of the femoral head(ONFH).Apoptosis of osteoblasts is considered as the main reason of ONFH at the molecular level.Glycogen synthase kinase 3β(GSK3β)is an important regulator of cellular differentiation and apoptosis pathway,which can modulate the balance between osteoblasts and osteoclasts.Several studies have reported about its function in osteoporosis,but little is known about it in osteonecrosis.In our study,lipopolysaccharide and methylprednisolone were utilized to establish a rat ONFH model.The phosphorylation of GSK3βSer-9 was decreased in the model.Western blotting examination ofβ-catenin,Bcl-2,Bax and caspase-3 revealed that the osteoblasts were apoptotic.In dexamethasone(Dex)-incubated primary osteoblasts,the expression profile of GSK3βphosphorylation and apoptotic factors were consistent with those in the rat ONFH model.To further investigate the regulation of osteonecrosis caused by GSK3β,the expression and function of GSK3βwere inhibited in Dex-incubated primary osteoblasts.The knockdown of GSK3βby siRNA decreased the expression of Bax and cleaved caspase-3,but increased Bcl-2 andβ-catenin.On the other hand,selective inhibition of GSK3βfunction by LiCl counteracted the activation of caspase-3 induced by Dex.Our work is the first study about the GSK3P phosphorylation in ONFH,and provides evidence for further therapeutic methods.
文摘Phosphorylation of protein is an important post-translational modification that enables activation of various enzymes and receptors included in signaling pathways. To reduce the cost of identifying phosphorylation site by laborious experiments, computational prediction of it has been actively studied. In this study, by adopting a new set of features and applying feature selection by Random Forest with grid search before training by Support Vector Machine, our method achieved better or comparable performance of phosphorylation site prediction for two different data sets.
基金Supported by the Italian Ministry of Health projects Ricerca Corrente1/17 and 21/17(Institute for Maternal and Child Health IRCCS Burlo Garofolo)
文摘BACKGROUND Intestinal inflammation and epithelial injury are the leading actors of inflammatory bowel disease(IBD),causing an excessive pro-inflammatory cytokines expression.Tristetraprolin(TTP),an mRNA binding protein,plays a role in regulating the inflammatory factors,recognizing specific sequences on the 3’untranslated region of cytokine mRNAs.TTP activity depends on its phosphorylation state:the unphosphorylated TTP degrades pro-inflammatory cytokine mRNAs;on the contrary,the phosphorylated TTP fails to destabilize mRNAs furthering their expression.The phospho-TTP forms a complex with the chaperone protein 14-3-3.This binding could be one of the factors that promote intestinal inflammation as a cause of disease progression.AIM To assess if TTP phosphorylation has a role in paediatric IBD.METHODS The study was carried out on a cohort of paediatric IBD patients.For each patient enrolled,a specimen of inflamed and non-inflamed colonic mucosa was collected.Furthermore,the experiments were conducted on macrophages differentiated from blood samples of the same patients.Macrophages from healthy donors’blood were used as controls.Co-immunoprecipitation assay and immunoblotting analyses were performed to observe the formation of the phospho-TTP/14-3-3 complex.In the same samples TNF-αexpression was also evaluated as major factor of the pro-inflammatory activity.RESULTS In this work we studied indirectly the phosphorylation of TTP through the binding with the chaperone protein 14-3-3.In inflamed and non-inflamed colon mucosa of IBD paediatric patients immunoblot assay demonstrated a higher expression of the TTP in inflamed samples respect to the non-inflamed;the coimmunoprecipitated 14-3-3 protein showed the same trend of expression.In the TNF-αgene expression analysis higher levels of the cytokine in inflamed tissues compared to controls were evident.The same experiments were conducted on macrophages from IBD paediatric patients and healthy controls.The immunoblot results demonstrated a high expression of both TTP and co-immunoprecipitated 14-4-3 protein in IBD-derived macrophages in comparison to healthy donors.TNF-αprotein levels from macrophages lysates showed the same trend of expression in favour of IBD paediatric patients compared to healthy controls.CONCLUSION In this work,for the first time,we describe a relation between phospho-TTP/14-3-3 complex and IBD.Indeed,a higher expression of TTP/14-3-3 was recorded in IBD samples in comparison to controls.
文摘Disturbances in constitutive nitric oxide synthase (cNOS) activation associated with H. pylori colonization of gastric mucosa are considered of major consequences in defining the extent of inflammatory involvement. As rapid changes in cNOS activation are linked to the enzyme phosphorylation at the specific Ser/Thr residues, we investigated the influence of H. pylori LPS and gastric hormone, ghrelin, on the processes of phosphorylation of these two critical sites in gastric mucosal cells. We show that the LPS-induced reduction in cNOS activity is reflected in the phosphorylation on Thr497, while the countering effect of ghrelin is associated with a rapid increase in cNOS phosphorylation on Ser1179. Further, we demonstrate that cNOS phosphorylation on Thr497 as well as Ser1179 displays dependence on PKCδ. However, while the LPS-induced suppression in cNOS activation shows reliance on the phosphorylation of PKCδ and PI3K on Ser, the effect of ghrelin is manifested by the increase in phosphorylation of PKCδ and PI3K on Tyr, as well as membrane translocation and phosphorylation of Akt on Ser493. Thus, our findings suggest that the LPS-induced suppression in cNOS activation is mediated by PKCδ-controlled phosphorylation of PI3K on Ser that interferes with the membrane recruitment of Akt and promotes cNOS phosphorylation on Thr497, and that ghrelin-elicited up-regulation in cNOS activation relies on the PKCδ-directed phosphorylation of PI3K on Tyr that stimulates the membrane localization of Akt and enhances cNOS phosphorylation on Ser1179.
文摘The signaling events underlying oral mucosal inflammatory responses to P. gingivalis and its key endotoxin, lipopolysaccharide (LPS), relay primarily on the LPS engagement of Toll-like receptor-4 (TLR4), and the activation of IκB-kinase complex (IKK) and mitogen-activated protein kinases (MAPKs that exert their control over transcription factors implicated in the regulation of iNOS and COX-2 proinflammatory genes expression). Since spleen tyrosine kinase (Syk) has emerged recently as a major amplifier in the production of proinflammatory mediators, we investigated the process of recruitment and interaction of Syk with TLR4 in salivary gland acinar cells in response to P. gingivalis LPS. Our findings revealed that stimulation of the acinar cells with the LPS leads to protein kinase Cδ (PKCδ)-mediated phosphorylation of Syk on Ser which results in its localization with the membrane associated TLR4 complex and the activation through phosphorylation on Tyr. Further, our results support the involvement of Syk in the amplification of transcription factors involved in the assembly and expression of transcription complexes associated with the induction in COX-2 and iNOS genes. Therefore, our data suggest that PKCδ is a primary linchpin affecting the Syk recruitment to the membrane localized TLR4, and hence affects the efficiency of the kinase activation and the magnitude of oral mucosal inflammatory response to P. gingivalis.
