Objective Kidney renal clear cell carcinoma(KIRC)is a common renal malignancy that has a poor prognosis.As a member of the F box family,cyclin F(CCNF)plays an important regulatory role in normal tissues and tumors.How...Objective Kidney renal clear cell carcinoma(KIRC)is a common renal malignancy that has a poor prognosis.As a member of the F box family,cyclin F(CCNF)plays an important regulatory role in normal tissues and tumors.However,the underlying mechanism by which CCNF promotes KIRC proliferation still remains unclear.Methods Bioinformatics methods were used to analyze The Cancer Genome Atlas(TCGA)database to obtain gene expression and clinical prognosis data.The CCK8 assay,EdU assay,and xenograft assay were used to detect cell proliferation.The cell senescence and potential mechanism were assessed by SA-β-gal staining,Western blotting,as well as ELISA.Results Our data showed that CCNF was highly expressed in KIRC patients.Meanwhile,downregulation of CCNF inhibited cell proliferation in vivo and in vitro.Further studies showed that the reduction of CCNF promoted cell senescence by decreasing cyclin-dependent kinase 1(CDK1),increasing the proinflammatory factors interleukin(IL)-6 and IL-8,and then enhancing the expression of p21 and p53.Conclusion We propose that the high expression of CCNF in KIRC may play a key role in tumorigenesis by regulating cell senescence.Therefore,CCNF shows promise as a new biomarker to predict the clinical prognosis of KIRC patients and as an effective therapeutic target.展开更多
BACKGROUND: The p25-activated cyclin-dependent protein kinase 5 (Cdk5) may induce neuronal cell death and cause the development of dementia following cerebral ischemia and reperfusion. OBJECTIVE: To observe change...BACKGROUND: The p25-activated cyclin-dependent protein kinase 5 (Cdk5) may induce neuronal cell death and cause the development of dementia following cerebral ischemia and reperfusion. OBJECTIVE: To observe changes in the expression of Cdk5 and p25 in hippocampal tissue of vascular dementia mice at different time points following cerebral ischemia and reperfusion. DESIGN, TIME AND SETTING: A randomized, controlled animal experiment was performed in the clinical trial center of Hebei Provincial People's Hospital between September 2007 and October 2008. MATERIALS: Cdk5 rabbit anti-mouse polyclonal antibody, p35 rabbit anti-mouse polyclonal antibody, and β-actin mouse monoclonal antibody were purchased from Santa Cruz Biotechnology, Inc., USA; horseradish peroxidase-labeled goat anti-rabbit IgG and horseradish peroxidase-labeled goat anti-mice IgG were offered by Beijing Zhongshan Geldenbridye Biotechnology Co.,Ltd., China; the protein quantitative kit was produced by Applygen Gene Technology Corp., Beijing, China; cDNA reverse transcription and PCR amplification reagents were products of TianGen& Biotech (Beijing) Co.,Ltd., China. METHODS: One hundred and sixty male Kunming mice were randomly divided into two groups: a sham-operated group (n = 65) and a model group (n = 95). Vascular dementia was induced with three periods of transient ischemia and reperfusion of the bilateral common carotid arteries. In the sham-operated group, the bilateral common carotid arteries were not blocked. MAIN OUTCOME MEASURES: Behavioral tests were done at four and six weeks post surgery. Pathological changes in the hippocampal CA1 region were observed with hematoxylin-eosin staining Cdk5 mRNA expression was examined by RT-PCR, and Western blots were used to evaluate Cdk5 and p25 expression. Learning and memory performance were assayed using the Morris water maze. RESULTS: Vascular dementia reduced learning and memory performance at 4 and 6 weeks post surgery. Vascular dementia also caused severe, time-dependent neuronal damage and death in the hippocampal CA1 region. Dementia induction also increased mRNA and protein expression of Cdk5 and p25 at both 4 and 6 weeks after surgery. CONCLUSION: Cdk5/p25 is involved in the development of vascular dementia in mice following cerebral ischemia and reperfusion.展开更多
It is known that human papillomavirus (HPV) infection can cause squamous cell neoplasms at several sites, such as cervix uteri carcinoma and oral squamous carcinoma. There is little information on the expression of ...It is known that human papillomavirus (HPV) infection can cause squamous cell neoplasms at several sites, such as cervix uteri carcinoma and oral squamous carcinoma. There is little information on the expression of HPV and its predictive markers in tumours of the major and minor salivary glands of the head and neck. We therefore assessed oral salivary gland neoplasms to identify associations between HPV and infection-related epidermal growth factor receptor (EGFR), cyclin-dependent kinase inhibitor 2A (CDKN2A/p16) and tumour protein p53 (TP53). Formalin-fixed, paraffin-embedded tissue samples from oral salivary gland carcinomas (n=51) and benign tumours (n=26) were analysed by polymerase chain reaction (PCR) analysis for several HPV species, including high-risk types 16 and 18. Evaluation of EGFR, CDKN2A, TP53 and cytomegalovirus (CMV) was performed by immunohistochemistry. Epstein-Barr virus (EBV) was evaluated by EBV-encoded RNA in situ hybridisation. We demonstrated that salivary gland tumours are not associated with HPV infection. The expression of EGFR, CDKN2A and TP53 may be associated with tumour pathology but is not induced by HPV. CMV and EBV were not detectable. In contrast to oral squamous cell carcinomas, HPV, CMV and EBV infections are not associated with malignant or benign neoplastic lesions of the salivary glands.展开更多
Background:The sensitivity of breast cancer cells to radiation is a key cause of locoregional recurrence after postoperative radiotherapy.Several studies have reported that microRNAs(miRNAs)are involved in the radiose...Background:The sensitivity of breast cancer cells to radiation is a key cause of locoregional recurrence after postoperative radiotherapy.Several studies have reported that microRNAs(miRNAs)are involved in the radiosensitivity of human breast cancer cells.One miRNA microarray study showed that miR-450b-5p was overexpressed 13.3-fold in patients with estrogen receptor–positive(ER^(+))and human epidermal growth factor receptor 2–negative(HER2−)breast cancer and no local relapse compared with local relapse patients.However,its underlying mechanism of action remains unknown.Methods:The predicted target mRNAs of miR-450b-5p were screened using the TargetScan,miRDB,and miRWalk databases.Western blotting,quantitative polymerase chain reaction,and dual-luciferase reporter assays explored the association between cyclindependent kinase 6(CDK6)and miR-450b-5p.The cell counting kit-8 assay and flow cytometry detected the proliferation of transfected MCF7 cells.Colony formation and xenograft tumors detected the radiosensitivity of the transfected MCF7 cells.Results:Bioinformatics analysis,Western blotting,quantitative polymerase chain reaction,and dual-luciferase reporter assays demonstrated that CDK6 was the target gene of miR-450b-5p.Furthermore,in vitro and in vivo experiments showed that miR-450b-5p inhibited MCF7 cell proliferation and cell cycle progression,increased the sensitizer enhancement ratio,and decreased the volume of xenograft tumors after irradiation by regulating CDK6.Conclusions:This study demonstrates that miR-450b-5p enhances the radiosensitivity of hormone receptor–positive(HR^(+))and HER2−breast cancer cells and elucidates its mechanism.miR-450b-5p may be considered a therapeutic target in HR^(+)and HER2−breast cancer treated with radiotherapy.展开更多
OBJECTIVE Previous studies have demonstrated acetylcholine muscarinic 4(M4) receptor regulates DARPP-32 phosphorylation at Thr75 in isolated medium spiny neurons(MSNs),indicating antagonistic mechanism with D1 depende...OBJECTIVE Previous studies have demonstrated acetylcholine muscarinic 4(M4) receptor regulates DARPP-32 phosphorylation at Thr75 in isolated medium spiny neurons(MSNs),indicating antagonistic mechanism with D1 dependent signal cascade,but the exact molecular mechanisms remain unclearly.In this study,we investigated the roles of M4 receptor in modulation D1 dependent signal to integrate striatal DA inputs in isolated MSNs.METHODS(1)Lentivirus technology was employed to genetically knock down the M4 receptor of MSNs;(2) Apomorphine(APO),acts as a dopamine receptor agonist,while SCH23390,acts as a selective antagonist for D1,were used to study the pharmacologically profiles with D1 receptor stimulation or blockade,respectively.Then the no subtype-selective muscarinic agonist oxotremorine M(OX) were used to show that mAchRs activation,in order to dissect the particular function of M4,a selective M4 antagonist,MT3 was used;(3) Intracellular cAMP production of MSNs was measured by using time resolved fluorescence resonance energy transfer detection method;(4) Laser confocal was used to explore the expression of M4 and D1 in MSNs;(5) Immunofluorescence cytochemistry and Western blotting were used to confirm the alteration of signaling molecular including P-CREB,DARPP-32 P-Thr34,DARPP-32 P-Thr75,cyclin-dependent kinase 5(CDK5) as wel as p25/35,which are involved in DA-dependent signaling modulations.RESULTS Firstly,TR-FRET assay revealed APO(10-2 mol·L^(-1))significantly increased the level of intracellular cAMP(vs control,n=3,P<0.01),also Western blotting results showed that APO(10-6 mol · L^(-1))increased DARPP-32 Thr34 phosphorylation(vs control,n=3,P<0.01),and these effect were reversed by D1 receptor antagonist SCH23390(vs APO,n=3,P<0.01).Interestingly,we confirmed that OX(10-6 mol · L^(-1)) down-regulated APO-induced DARPP-32 Thr34 phosphorylation(vs APO,n=3,P<0.01),due to its effects on DARPP-32 phosphorylation at Thr75.The results presented the antagonistic mechanism of mAchRs stimulation with D1 dependent signal cascade in MSNs.Meanwhile,OX(10-7,10-6 and10^(-5) mol·L^(-1)) stimulated DARPP-32 phosphorylation at Thr75,and simultaneously up regulated P25/35 and CDK5 activity(vs control,n=3,P<0.01) by using Western blotting assay.Furthermore,roscovitine(10^(-5) mol · L^(-1)),acts as a CDK5 inhibitor,suppressed CDK5 activity(vs control,n=10,P<0.01),and fully inhibited OX-induced DARPP-32 Thr75 phosphorylation(vs OX,n=10,P<0.01).More important,pretreated with roscovitine(10^(-5) mol·L^(-1)),the effect of APO on DARPP-32 Thr34 phosphorylation was potentiated(vs APO,n=3,P<0.05).The result presented CDK5 is required in suppression of APO on DARPP-32 Thr34 phosphorylation mediated through mAchRs stimulation.In addition,laser confocal results showed that the CDK5 up-regulation was mostly confined to MSNs co-expressing M4,which means that M4 participated in CDK5-mediated phosphorylation of DARPP-32 at Thr75.Consistently,immunofluorescence and Western blotting results confirmed that both genetic knockdown and pharmacologic inhibition of M4 receptors with MT3(10-7 mol · L^(-1)) down-regulated the OX-induced the expression of CDK5(vs OX,n=3,P<0.01) and P25/35(vs OX,n=3,P<0.01)in isolated MSNs.CONCLUSION M4 receptor may play an important role in antagonistic regulation D1 dependent signaling,in which CDK5 is required for suppressing D1-DARPP-32 Thr34 phosphorylation in isolated medium spiny neurons.展开更多
Abemaciclib (Verzerio<span style="white-space:nowrap;"><sup><span style="font-family:Verdana, Helvetica, Arial;white-space:normal;background-color:#FFFFFF;">®</span>...Abemaciclib (Verzerio<span style="white-space:nowrap;"><sup><span style="font-family:Verdana, Helvetica, Arial;white-space:normal;background-color:#FFFFFF;">®</span></sup></span>) is a cell cycle inhibitor of both CDK4 and CDK6. In 2017, abemaciclib was approved by the Food and Drug Administration (FDA) and, in 2018 by the European Medicines Agency (EMA) for the treatment of postmenopausal women with hormone receptor positive (HR<sup>+</sup>), human epidermal growth factor receptor 2 negative (HER2<sup><span style="white-space:nowrap;"><sup><span style="white-space:nowrap;">−</span></sup></span></sup>) advanced breast cancer. In this mini-review, we provide a series of information for respectively their targets and its selectivity, results on preclinical trial, clinical phase I, II and III trials, and some perspectives. We also describe the batch and flow steps used for the synthesis of this cancer drug.展开更多
BACKGROUND Pathological complete response(pCR) is rare in hormone receptor-positive(HR+)HER2-negative breast cancer(BC) treated with either endocrine therapy(ET) or chemotherapy. Radical resection of locoregional rela...BACKGROUND Pathological complete response(pCR) is rare in hormone receptor-positive(HR+)HER2-negative breast cancer(BC) treated with either endocrine therapy(ET) or chemotherapy. Radical resection of locoregional relapse, although potentially curative in some cases, is challenging when the tumor invades critical structures.The oral cyclin-dependent kinase 4/6 inhibitor palbociclib in combination with ET has obtained a significant increase in objective response rates and progression-free survival in patients with advanced BC and is now being evaluated in the neoadjuvant setting. We present a clinical case of a patient with an inoperable locoregional relapse of HR+ HER2-negative BC who experienced p CR after treatment with palbociclib.CASE SUMMARY We report the clinical case of a 60-year-old patient who presented with an inoperable locoregional relapse of HR+, HER2-negative BC 10 years after the diagnosis of the primary tumor. During a routine follow-up visit, breast magnetic resonance imaging and positron emission tomography/computed tomography revealed a 4-cm lesion in the right subclavicular region, infiltrating the chest wall and extending to the subclavian vessels, but without bone or visceral involvement. Treatment was begun with palbociclib plus letrozole, converting the disease to operability over a period of 6 mo. Surgery was performed and a p CR achieved. Of note, during treatment the patient experienced a very uncommon toxicity characterized by burning tongue and glossodynia associated with dysgeusia, paresthesia, dysesthesia, and xerostomia. A reduction in the dose of palbociclib did not provide relief and treatment with the inhibitor was thus discontinued, resolving the tongue symptoms. Laboratory exams were unremarkable. Given that this was a late relapse, the tumor was classified asendocrine-sensitive, a condition associated with high sensitivity to palbociclib.CONCLUSION This case highlights the potential of the cyclin-dependent kinase 4/6 inhibitor plus ET combination to achieve pCR in locoregional relapse of BC, enabling surgical resection of a lesion initially considered inoperable.展开更多
This study was designed to investigate the relationship of the expression of cyclin-dependent kinases (CDKs) with theeffects of all-trans retinoic acid (ATRA) on the proliferation of HL-cells. HL-60 cells were treated...This study was designed to investigate the relationship of the expression of cyclin-dependent kinases (CDKs) with theeffects of all-trans retinoic acid (ATRA) on the proliferation of HL-cells. HL-60 cells were treated with ATRA for 1-4 d. Then thecapacity of DNA Synthesis was evaluated with 3H-TdR incorporation and the expression of cyclin E, cyclin D, CDK2 and CDK4protein determined with immunocytochemical staining. In addition, the expression Of CDC2, CDK2 and CDK4 mRNA was deter-mined with in situ hybridization. It was found that ATRA suppressed the proliferation of HL-60 cells and decreased their capacityof DNA synthesis to result in a down-regulation of the expression of cyclin E, cyclin D and CDC2 without comcomittant suppressionon the expression of CDK2 and CDK4. It is concluded that the effects of ATRA on the proliferation of HL-60 cells may be relatedto the down-regulation of the expression of cyclin E, cyclin D and CDC2.展开更多
Hepatocellular carcinoma (HCC) is one of the most common human cancers, and its incidence is still increasing in many countries. The prognosis of HCC patients remains poor, and identification of useful molecular pro...Hepatocellular carcinoma (HCC) is one of the most common human cancers, and its incidence is still increasing in many countries. The prognosis of HCC patients remains poor, and identification of useful molecular prognostic markers is required. Many recent studies have shown that functional alterations of cellcycle regulators can be observed in HCC. Among the various types of cell-cycle regulators, p16 and p27 are frequently inactivated in HCC and are considered to be potent tumor suppressors, p16, a G1-specific cell-cycle inhibitor that prevents the association of cyclindependent kinase (CDK) 4 and CDK6 with cyclin DI, is frequently inactivated in HCC via CpG methylation of its promoter region, p16 may be involved in the early steps of hepatocarcinogenesis, since p16 gene methylation has been detected in subsets of pre-neoplastic liver cirrhosis patients, p27, a negative regulator of the G1-S phase transition through inhibition of the kinase activities of Cdk2/cyclin A and Cdk2/cyclin E complexes, is now considered to be an adverse prognostic factor in HCC. In some cases of HCC with increased cell proliferation, p27 is overexpressed but inactivated by sequestration into cyclin D1-CDK4-containing complexes. Since loss of p16 is closely related to functional inactivation of p27 in HCC, investigating both p16 and p27 may be useful for precise prognostic predictions in individuals with HCC.展开更多
Hepatocellular carcinoma (HCC) is the leading cause of cancer death worldwide;nevertheless, currenttherapeutic options are limited or ineffective for many patients. Therefore, elucidation of molecular mechanisms inHCC...Hepatocellular carcinoma (HCC) is the leading cause of cancer death worldwide;nevertheless, currenttherapeutic options are limited or ineffective for many patients. Therefore, elucidation of molecular mechanisms inHCC biology could yield important insights for the intervention of novel therapies. Recently, various studies havereported dysregulation of long non-coding RNAs (lncRNAs) in the initiation and progression of HCC, including H19;however, the biological function of H19 in HCC remains unclear. Here, we show that knockdown of H19 disruptedHCC cell growth, impaired the G1-to-S phase transition, and promoted apoptosis, while overexpression of H19yielded the opposite results. Screening for expression of cell cycle-related genes revealed a significant downregulationof CDK6 at both RNA and protein levels upon H19 suppression. Bioinformatic analysis of the H19 sequence and the3′ untranslated region (3′ UTR) of CDK6 transcripts showed several binding sites for microRNA-107 (miR-107), andthe dual luciferase reporter assay confirmed their direct interaction with miR-107. Consistently, blockage of miR-107activity alleviated the growth suppression phenotypes induced by H19 downregulation, suggesting that H19 serves asa molecular sponge for miR-107 to promote CDK6 expression and cell cycle progression. Together, this studydemonstrates a mechanistic function of H19 in driving the proliferation of HCC cells and suggests H19 suppressionas a novel approach for HCC treatment.展开更多
Objective Post-stroke cognitive impairment(PSCI)develops in approximately one-third of stroke survivors and is associated with ingravescence.Nonetheless,the biochemical mechanisms underlying PSCI remain unclear.The st...Objective Post-stroke cognitive impairment(PSCI)develops in approximately one-third of stroke survivors and is associated with ingravescence.Nonetheless,the biochemical mechanisms underlying PSCI remain unclear.The study aimed to establish an ischemic mouse model by means of transient unilateral middle cerebral artery occlusions(MCAOs)and to explore the biochemical mechanisms of p25/cyclin-dependent kinase 5(CDK5)-mediated tau hyperphosphorylation on the PSCI behavior.Methods Cognitive behavior was investigated,followed by the detection of tau hyperphosphorylation,mobilization,activation of kinases and/or inhibition of phosphatases in the lateral and contralateral cerebrum of mice following ischemia in MACO mice.Finally,we treated HEK293/tau cells with oxygen-glucose deprivation(OGD)and a CDK5 inhibitor(Roscovitine)or a GSK3βinhibitor(LiCl)to the roles of CDK5 and GSK3βin mediating ischemia-reperfusion-induced tau phosphorylation.Results Ischemia induced cognitive impairments within 2 months,as well as causing tau hyperphosphorylation and its localization to neuronal somata in both ipsilateral and contralateral cerebra.Furthermore,p25 that promotes CDK5 hyperactivation had significantly higher expression in the mice with MCAO than in the shamoperation(control)group,while the expression levels of protein phosphatase 2(PP2A)and the phosphorylation level at Tyr307 were comparable between the two groups.In addition,the CDK5 inhibitor rescued tau from hyperphosphorylation induced by OGD.Conclusion These findings demonstrate that upregulation of CDK5 mediates tau hyperphosphorylation and localization in both ipsilateral and contralateral cerebra,contributing to the pathogenesis of PSCI.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 investigate the mechanism of cell cyclin-dependent kinase (KDM5B), a key enzyme driving all cell cycle transitions, promoting HCC progression and metastasis. Methods: The expression of KDM5B in normal li...Objective: To investigate the mechanism of cell cyclin-dependent kinase (KDM5B), a key enzyme driving all cell cycle transitions, promoting HCC progression and metastasis. Methods: The expression of KDM5B in normal liver, HCC and its adjacent tissues was analyzed by RT-PCR and IHC. Lentivirus transfection method was used to construct stable cell lines with KDM5B overexpression and down-regulation, and the role of KDM5B in HCC migration and invasion was detected at cell level and animal level. Western blotting and Transwell experiments were performed to verify the effect of KDM5B and/or CCR2 inhibitors on HCC progression and metastasis by using liver orthotopic transplantation tumor model and immunofluorescence methods. Results: RT-PCR showed that the expression level of KDM5B in HCC was significantly higher than that in adjacent tissues, and the increase of KDM5B was relatively significant. Upregulation of KDM5B in nude mouse liver orthotopic transplantation tumor model can promote the incidence of lung metastasis and shorten the survival time of nude mice, whereas upregulation of KDM5B can reduce the incidence of lung metastasis and prolong the survival time of nude mice. Conclusion: This study clarified the expression of KDM5B in HCC and its function in promoting HCC migration, invasion and metastasis. The molecular mechanism of KDM5B promoting HCC metastasis was revealed, providing a potential therapeutic target for HCC.展开更多
基金supported by the National Natural Science Foundation of China(No.81874148 and No.82203142).
文摘Objective Kidney renal clear cell carcinoma(KIRC)is a common renal malignancy that has a poor prognosis.As a member of the F box family,cyclin F(CCNF)plays an important regulatory role in normal tissues and tumors.However,the underlying mechanism by which CCNF promotes KIRC proliferation still remains unclear.Methods Bioinformatics methods were used to analyze The Cancer Genome Atlas(TCGA)database to obtain gene expression and clinical prognosis data.The CCK8 assay,EdU assay,and xenograft assay were used to detect cell proliferation.The cell senescence and potential mechanism were assessed by SA-β-gal staining,Western blotting,as well as ELISA.Results Our data showed that CCNF was highly expressed in KIRC patients.Meanwhile,downregulation of CCNF inhibited cell proliferation in vivo and in vitro.Further studies showed that the reduction of CCNF promoted cell senescence by decreasing cyclin-dependent kinase 1(CDK1),increasing the proinflammatory factors interleukin(IL)-6 and IL-8,and then enhancing the expression of p21 and p53.Conclusion We propose that the high expression of CCNF in KIRC may play a key role in tumorigenesis by regulating cell senescence.Therefore,CCNF shows promise as a new biomarker to predict the clinical prognosis of KIRC patients and as an effective therapeutic target.
文摘BACKGROUND: The p25-activated cyclin-dependent protein kinase 5 (Cdk5) may induce neuronal cell death and cause the development of dementia following cerebral ischemia and reperfusion. OBJECTIVE: To observe changes in the expression of Cdk5 and p25 in hippocampal tissue of vascular dementia mice at different time points following cerebral ischemia and reperfusion. DESIGN, TIME AND SETTING: A randomized, controlled animal experiment was performed in the clinical trial center of Hebei Provincial People's Hospital between September 2007 and October 2008. MATERIALS: Cdk5 rabbit anti-mouse polyclonal antibody, p35 rabbit anti-mouse polyclonal antibody, and β-actin mouse monoclonal antibody were purchased from Santa Cruz Biotechnology, Inc., USA; horseradish peroxidase-labeled goat anti-rabbit IgG and horseradish peroxidase-labeled goat anti-mice IgG were offered by Beijing Zhongshan Geldenbridye Biotechnology Co.,Ltd., China; the protein quantitative kit was produced by Applygen Gene Technology Corp., Beijing, China; cDNA reverse transcription and PCR amplification reagents were products of TianGen& Biotech (Beijing) Co.,Ltd., China. METHODS: One hundred and sixty male Kunming mice were randomly divided into two groups: a sham-operated group (n = 65) and a model group (n = 95). Vascular dementia was induced with three periods of transient ischemia and reperfusion of the bilateral common carotid arteries. In the sham-operated group, the bilateral common carotid arteries were not blocked. MAIN OUTCOME MEASURES: Behavioral tests were done at four and six weeks post surgery. Pathological changes in the hippocampal CA1 region were observed with hematoxylin-eosin staining Cdk5 mRNA expression was examined by RT-PCR, and Western blots were used to evaluate Cdk5 and p25 expression. Learning and memory performance were assayed using the Morris water maze. RESULTS: Vascular dementia reduced learning and memory performance at 4 and 6 weeks post surgery. Vascular dementia also caused severe, time-dependent neuronal damage and death in the hippocampal CA1 region. Dementia induction also increased mRNA and protein expression of Cdk5 and p25 at both 4 and 6 weeks after surgery. CONCLUSION: Cdk5/p25 is involved in the development of vascular dementia in mice following cerebral ischemia and reperfusion.
文摘It is known that human papillomavirus (HPV) infection can cause squamous cell neoplasms at several sites, such as cervix uteri carcinoma and oral squamous carcinoma. There is little information on the expression of HPV and its predictive markers in tumours of the major and minor salivary glands of the head and neck. We therefore assessed oral salivary gland neoplasms to identify associations between HPV and infection-related epidermal growth factor receptor (EGFR), cyclin-dependent kinase inhibitor 2A (CDKN2A/p16) and tumour protein p53 (TP53). Formalin-fixed, paraffin-embedded tissue samples from oral salivary gland carcinomas (n=51) and benign tumours (n=26) were analysed by polymerase chain reaction (PCR) analysis for several HPV species, including high-risk types 16 and 18. Evaluation of EGFR, CDKN2A, TP53 and cytomegalovirus (CMV) was performed by immunohistochemistry. Epstein-Barr virus (EBV) was evaluated by EBV-encoded RNA in situ hybridisation. We demonstrated that salivary gland tumours are not associated with HPV infection. The expression of EGFR, CDKN2A and TP53 may be associated with tumour pathology but is not induced by HPV. CMV and EBV were not detectable. In contrast to oral squamous cell carcinomas, HPV, CMV and EBV infections are not associated with malignant or benign neoplastic lesions of the salivary glands.
文摘Background:The sensitivity of breast cancer cells to radiation is a key cause of locoregional recurrence after postoperative radiotherapy.Several studies have reported that microRNAs(miRNAs)are involved in the radiosensitivity of human breast cancer cells.One miRNA microarray study showed that miR-450b-5p was overexpressed 13.3-fold in patients with estrogen receptor–positive(ER^(+))and human epidermal growth factor receptor 2–negative(HER2−)breast cancer and no local relapse compared with local relapse patients.However,its underlying mechanism of action remains unknown.Methods:The predicted target mRNAs of miR-450b-5p were screened using the TargetScan,miRDB,and miRWalk databases.Western blotting,quantitative polymerase chain reaction,and dual-luciferase reporter assays explored the association between cyclindependent kinase 6(CDK6)and miR-450b-5p.The cell counting kit-8 assay and flow cytometry detected the proliferation of transfected MCF7 cells.Colony formation and xenograft tumors detected the radiosensitivity of the transfected MCF7 cells.Results:Bioinformatics analysis,Western blotting,quantitative polymerase chain reaction,and dual-luciferase reporter assays demonstrated that CDK6 was the target gene of miR-450b-5p.Furthermore,in vitro and in vivo experiments showed that miR-450b-5p inhibited MCF7 cell proliferation and cell cycle progression,increased the sensitizer enhancement ratio,and decreased the volume of xenograft tumors after irradiation by regulating CDK6.Conclusions:This study demonstrates that miR-450b-5p enhances the radiosensitivity of hormone receptor–positive(HR^(+))and HER2−breast cancer cells and elucidates its mechanism.miR-450b-5p may be considered a therapeutic target in HR^(+)and HER2−breast cancer treated with radiotherapy.
文摘OBJECTIVE Previous studies have demonstrated acetylcholine muscarinic 4(M4) receptor regulates DARPP-32 phosphorylation at Thr75 in isolated medium spiny neurons(MSNs),indicating antagonistic mechanism with D1 dependent signal cascade,but the exact molecular mechanisms remain unclearly.In this study,we investigated the roles of M4 receptor in modulation D1 dependent signal to integrate striatal DA inputs in isolated MSNs.METHODS(1)Lentivirus technology was employed to genetically knock down the M4 receptor of MSNs;(2) Apomorphine(APO),acts as a dopamine receptor agonist,while SCH23390,acts as a selective antagonist for D1,were used to study the pharmacologically profiles with D1 receptor stimulation or blockade,respectively.Then the no subtype-selective muscarinic agonist oxotremorine M(OX) were used to show that mAchRs activation,in order to dissect the particular function of M4,a selective M4 antagonist,MT3 was used;(3) Intracellular cAMP production of MSNs was measured by using time resolved fluorescence resonance energy transfer detection method;(4) Laser confocal was used to explore the expression of M4 and D1 in MSNs;(5) Immunofluorescence cytochemistry and Western blotting were used to confirm the alteration of signaling molecular including P-CREB,DARPP-32 P-Thr34,DARPP-32 P-Thr75,cyclin-dependent kinase 5(CDK5) as wel as p25/35,which are involved in DA-dependent signaling modulations.RESULTS Firstly,TR-FRET assay revealed APO(10-2 mol·L^(-1))significantly increased the level of intracellular cAMP(vs control,n=3,P<0.01),also Western blotting results showed that APO(10-6 mol · L^(-1))increased DARPP-32 Thr34 phosphorylation(vs control,n=3,P<0.01),and these effect were reversed by D1 receptor antagonist SCH23390(vs APO,n=3,P<0.01).Interestingly,we confirmed that OX(10-6 mol · L^(-1)) down-regulated APO-induced DARPP-32 Thr34 phosphorylation(vs APO,n=3,P<0.01),due to its effects on DARPP-32 phosphorylation at Thr75.The results presented the antagonistic mechanism of mAchRs stimulation with D1 dependent signal cascade in MSNs.Meanwhile,OX(10-7,10-6 and10^(-5) mol·L^(-1)) stimulated DARPP-32 phosphorylation at Thr75,and simultaneously up regulated P25/35 and CDK5 activity(vs control,n=3,P<0.01) by using Western blotting assay.Furthermore,roscovitine(10^(-5) mol · L^(-1)),acts as a CDK5 inhibitor,suppressed CDK5 activity(vs control,n=10,P<0.01),and fully inhibited OX-induced DARPP-32 Thr75 phosphorylation(vs OX,n=10,P<0.01).More important,pretreated with roscovitine(10^(-5) mol·L^(-1)),the effect of APO on DARPP-32 Thr34 phosphorylation was potentiated(vs APO,n=3,P<0.05).The result presented CDK5 is required in suppression of APO on DARPP-32 Thr34 phosphorylation mediated through mAchRs stimulation.In addition,laser confocal results showed that the CDK5 up-regulation was mostly confined to MSNs co-expressing M4,which means that M4 participated in CDK5-mediated phosphorylation of DARPP-32 at Thr75.Consistently,immunofluorescence and Western blotting results confirmed that both genetic knockdown and pharmacologic inhibition of M4 receptors with MT3(10-7 mol · L^(-1)) down-regulated the OX-induced the expression of CDK5(vs OX,n=3,P<0.01) and P25/35(vs OX,n=3,P<0.01)in isolated MSNs.CONCLUSION M4 receptor may play an important role in antagonistic regulation D1 dependent signaling,in which CDK5 is required for suppressing D1-DARPP-32 Thr34 phosphorylation in isolated medium spiny neurons.
文摘Abemaciclib (Verzerio<span style="white-space:nowrap;"><sup><span style="font-family:Verdana, Helvetica, Arial;white-space:normal;background-color:#FFFFFF;">®</span></sup></span>) is a cell cycle inhibitor of both CDK4 and CDK6. In 2017, abemaciclib was approved by the Food and Drug Administration (FDA) and, in 2018 by the European Medicines Agency (EMA) for the treatment of postmenopausal women with hormone receptor positive (HR<sup>+</sup>), human epidermal growth factor receptor 2 negative (HER2<sup><span style="white-space:nowrap;"><sup><span style="white-space:nowrap;">−</span></sup></span></sup>) advanced breast cancer. In this mini-review, we provide a series of information for respectively their targets and its selectivity, results on preclinical trial, clinical phase I, II and III trials, and some perspectives. We also describe the batch and flow steps used for the synthesis of this cancer drug.
文摘BACKGROUND Pathological complete response(pCR) is rare in hormone receptor-positive(HR+)HER2-negative breast cancer(BC) treated with either endocrine therapy(ET) or chemotherapy. Radical resection of locoregional relapse, although potentially curative in some cases, is challenging when the tumor invades critical structures.The oral cyclin-dependent kinase 4/6 inhibitor palbociclib in combination with ET has obtained a significant increase in objective response rates and progression-free survival in patients with advanced BC and is now being evaluated in the neoadjuvant setting. We present a clinical case of a patient with an inoperable locoregional relapse of HR+ HER2-negative BC who experienced p CR after treatment with palbociclib.CASE SUMMARY We report the clinical case of a 60-year-old patient who presented with an inoperable locoregional relapse of HR+, HER2-negative BC 10 years after the diagnosis of the primary tumor. During a routine follow-up visit, breast magnetic resonance imaging and positron emission tomography/computed tomography revealed a 4-cm lesion in the right subclavicular region, infiltrating the chest wall and extending to the subclavian vessels, but without bone or visceral involvement. Treatment was begun with palbociclib plus letrozole, converting the disease to operability over a period of 6 mo. Surgery was performed and a p CR achieved. Of note, during treatment the patient experienced a very uncommon toxicity characterized by burning tongue and glossodynia associated with dysgeusia, paresthesia, dysesthesia, and xerostomia. A reduction in the dose of palbociclib did not provide relief and treatment with the inhibitor was thus discontinued, resolving the tongue symptoms. Laboratory exams were unremarkable. Given that this was a late relapse, the tumor was classified asendocrine-sensitive, a condition associated with high sensitivity to palbociclib.CONCLUSION This case highlights the potential of the cyclin-dependent kinase 4/6 inhibitor plus ET combination to achieve pCR in locoregional relapse of BC, enabling surgical resection of a lesion initially considered inoperable.
文摘This study was designed to investigate the relationship of the expression of cyclin-dependent kinases (CDKs) with theeffects of all-trans retinoic acid (ATRA) on the proliferation of HL-cells. HL-60 cells were treated with ATRA for 1-4 d. Then thecapacity of DNA Synthesis was evaluated with 3H-TdR incorporation and the expression of cyclin E, cyclin D, CDK2 and CDK4protein determined with immunocytochemical staining. In addition, the expression Of CDC2, CDK2 and CDK4 mRNA was deter-mined with in situ hybridization. It was found that ATRA suppressed the proliferation of HL-60 cells and decreased their capacityof DNA synthesis to result in a down-regulation of the expression of cyclin E, cyclin D and CDC2 without comcomittant suppressionon the expression of CDK2 and CDK4. It is concluded that the effects of ATRA on the proliferation of HL-60 cells may be relatedto the down-regulation of the expression of cyclin E, cyclin D and CDC2.
文摘Hepatocellular carcinoma (HCC) is one of the most common human cancers, and its incidence is still increasing in many countries. The prognosis of HCC patients remains poor, and identification of useful molecular prognostic markers is required. Many recent studies have shown that functional alterations of cellcycle regulators can be observed in HCC. Among the various types of cell-cycle regulators, p16 and p27 are frequently inactivated in HCC and are considered to be potent tumor suppressors, p16, a G1-specific cell-cycle inhibitor that prevents the association of cyclindependent kinase (CDK) 4 and CDK6 with cyclin DI, is frequently inactivated in HCC via CpG methylation of its promoter region, p16 may be involved in the early steps of hepatocarcinogenesis, since p16 gene methylation has been detected in subsets of pre-neoplastic liver cirrhosis patients, p27, a negative regulator of the G1-S phase transition through inhibition of the kinase activities of Cdk2/cyclin A and Cdk2/cyclin E complexes, is now considered to be an adverse prognostic factor in HCC. In some cases of HCC with increased cell proliferation, p27 is overexpressed but inactivated by sequestration into cyclin D1-CDK4-containing complexes. Since loss of p16 is closely related to functional inactivation of p27 in HCC, investigating both p16 and p27 may be useful for precise prognostic predictions in individuals with HCC.
基金financially supported by Thailand Science Research and Innovation Fund Chulalongkorn University(CU_FRB65_hea(46)_053_30_34)Ratchadapiseksompotch Fund,Faculty of Medicine,Chulalongkorn University(Grant No.RA 66/017)+1 种基金Thailand Research Fund(TRF)Senior Research Scholar(Grant No.RTA6280004)the Center of Excellence in Hepatitis and Liver Cancer,Faculty of Medicine,Chulalongkorn University.
文摘Hepatocellular carcinoma (HCC) is the leading cause of cancer death worldwide;nevertheless, currenttherapeutic options are limited or ineffective for many patients. Therefore, elucidation of molecular mechanisms inHCC biology could yield important insights for the intervention of novel therapies. Recently, various studies havereported dysregulation of long non-coding RNAs (lncRNAs) in the initiation and progression of HCC, including H19;however, the biological function of H19 in HCC remains unclear. Here, we show that knockdown of H19 disruptedHCC cell growth, impaired the G1-to-S phase transition, and promoted apoptosis, while overexpression of H19yielded the opposite results. Screening for expression of cell cycle-related genes revealed a significant downregulationof CDK6 at both RNA and protein levels upon H19 suppression. Bioinformatic analysis of the H19 sequence and the3′ untranslated region (3′ UTR) of CDK6 transcripts showed several binding sites for microRNA-107 (miR-107), andthe dual luciferase reporter assay confirmed their direct interaction with miR-107. Consistently, blockage of miR-107activity alleviated the growth suppression phenotypes induced by H19 downregulation, suggesting that H19 serves asa molecular sponge for miR-107 to promote CDK6 expression and cell cycle progression. Together, this studydemonstrates a mechanistic function of H19 in driving the proliferation of HCC cells and suggests H19 suppressionas a novel approach for HCC treatment.
基金the National Natural Science Foundation of China(No.31800851)Natural Science Foundation of Hubei Province(No.2022CFB456)The Research Fund of Jianghan University(No.08210011).
文摘Objective Post-stroke cognitive impairment(PSCI)develops in approximately one-third of stroke survivors and is associated with ingravescence.Nonetheless,the biochemical mechanisms underlying PSCI remain unclear.The study aimed to establish an ischemic mouse model by means of transient unilateral middle cerebral artery occlusions(MCAOs)and to explore the biochemical mechanisms of p25/cyclin-dependent kinase 5(CDK5)-mediated tau hyperphosphorylation on the PSCI behavior.Methods Cognitive behavior was investigated,followed by the detection of tau hyperphosphorylation,mobilization,activation of kinases and/or inhibition of phosphatases in the lateral and contralateral cerebrum of mice following ischemia in MACO mice.Finally,we treated HEK293/tau cells with oxygen-glucose deprivation(OGD)and a CDK5 inhibitor(Roscovitine)or a GSK3βinhibitor(LiCl)to the roles of CDK5 and GSK3βin mediating ischemia-reperfusion-induced tau phosphorylation.Results Ischemia induced cognitive impairments within 2 months,as well as causing tau hyperphosphorylation and its localization to neuronal somata in both ipsilateral and contralateral cerebra.Furthermore,p25 that promotes CDK5 hyperactivation had significantly higher expression in the mice with MCAO than in the shamoperation(control)group,while the expression levels of protein phosphatase 2(PP2A)and the phosphorylation level at Tyr307 were comparable between the two groups.In addition,the CDK5 inhibitor rescued tau from hyperphosphorylation induced by OGD.Conclusion These findings demonstrate that upregulation of CDK5 mediates tau hyperphosphorylation and localization in both ipsilateral and contralateral cerebra,contributing to the pathogenesis of PSCI.
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.
文摘Objective: To investigate the mechanism of cell cyclin-dependent kinase (KDM5B), a key enzyme driving all cell cycle transitions, promoting HCC progression and metastasis. Methods: The expression of KDM5B in normal liver, HCC and its adjacent tissues was analyzed by RT-PCR and IHC. Lentivirus transfection method was used to construct stable cell lines with KDM5B overexpression and down-regulation, and the role of KDM5B in HCC migration and invasion was detected at cell level and animal level. Western blotting and Transwell experiments were performed to verify the effect of KDM5B and/or CCR2 inhibitors on HCC progression and metastasis by using liver orthotopic transplantation tumor model and immunofluorescence methods. Results: RT-PCR showed that the expression level of KDM5B in HCC was significantly higher than that in adjacent tissues, and the increase of KDM5B was relatively significant. Upregulation of KDM5B in nude mouse liver orthotopic transplantation tumor model can promote the incidence of lung metastasis and shorten the survival time of nude mice, whereas upregulation of KDM5B can reduce the incidence of lung metastasis and prolong the survival time of nude mice. Conclusion: This study clarified the expression of KDM5B in HCC and its function in promoting HCC migration, invasion and metastasis. The molecular mechanism of KDM5B promoting HCC metastasis was revealed, providing a potential therapeutic target for HCC.