Long-term trends in the burden of colorectal cancer in Europe over three decades: a joinpoint regression and age-period-cohort analysis.

Background: The burden of colorectal cancer (CRC) in Europe is at a high level, but the epidemiological features have not yet been systematically studied. This study aimed to provide a timely and reliable assessment of the burden and trends of CRC in Europe to provide a scientific basis for its prevention and treatment. Methods: We analyzed data on CRC in 44 European countries between 1990 and 2019 from the Global Burden of Disease study (GBD) 2019. In addition, the joinpoint regression model was applied to reflect temporal trends. The age-period-cohort model was constructed to explore age, period, and birth cohort effects that influence the risk of morbidity and mortality. Results: In Europe, new cases, disability-adjusted life years (DALYs) and deaths of CRC rose by 70.01%, 22.88% and 38.04% from 1990 to 2019, respectively. The age-standardized incidence rate (ASIR) has increased, while age-standardized DALY rate and age-standardized mortality rate (ASMR) have declined. We found that men experienced a significantly higher CRC burden than women. Age-period-cohort analysis showed that the risk of incidence and mortality increased with age and time; and it was lower in the later-born cohort than the earlier-born cohort. Conclusion: ASIR for CRC in Europe generally trended upwards from 1990 to 2019, stabilizing in recent years but still at a high level. CRC burden varied considerably in different countries. There was a pronounced gender difference in CRC burden, and middle-aged and older men should be a priority population for CRC prevention and treatment.

Different Sedation Strategies in Older Patients Receiving Spinal Anesthesia for Hip Surgery on Postoperative Delirium: A Randomized Clinical Trial.

Background: Postoperative delirium (POD) is of great concern as a complication of surgery in older adult patients. Sedation strategies influence the development of POD. This study compared how sedation strategies administered during spinal anesthesia influenced POD in patients aged ≥65 years undergoing elective surgery for hip fracture repair. Patients and Methods: A randomized clinical trial was conducted from 1 August 2021 to 30 June 2022 at a single academic medical center. Two hundred and twenty-six patients were randomly divided into four groups: lighter sedation with propofol (LP), heavier sedation with propofol (HP), lighter sedation with dexmedetomidine (LD), and heavier sedation with dexmedetomidine (HD). The incidence of delirium was the primary outcome and was assessed daily by the blinded Confusion Assessment Method. Results: There was a significant association between dexmedetomidine (LD+HD group) and a lower incidence of delirium (11.9% [13/109] vs the propofol group (23.6% [26/110]; Risk ratio, 0.51; 95% CI, 0.274 to 0.929; p=0.024). In the propofol group, heavier sedation had a higher rate of POD (32.7% [18/55] vs the lighter sedation group (14.5% [8/55]; Risk ratio, 2.25; 95% CI, 1.069 to 4.736; p=0.025). Conclusion: Dexmedetomidine was associated with a lower incidence of delirium than that with propofol among older patients with hip fractures. In patients that received propofol, heavier sedation was associated with high incidence of POD.

Effect of propofol-based total intravenous anaesthesia on postoperative cognitive function and sleep quality in elderly patients.

PURPOSE: This study aimed to observe the effect of propofol-based total intravenous anaesthesia (TIVA) on postoperative cognitive function and sleep quality in elderly patients. METHODS: From August 2019 to August 2020, 130 cases of elderly patients who underwent abdominal surgery in The Third Hospital of Hebei Medical University were enrolled in this study. All participants were randomly divided into TIVA group (n = 65, receiving propofol-based TIVA) and control group (n = 65, receiving inhaled of sevoflurane anaesthesia). Intra-operative indicators were recorded in both groups. The cognitive function, sleep quality, urine melatonin sulphate, free cortisol, S-100 ß protein and interleukin-6 (IL-6) levels were compared at different times. RESULTS: On 1, 3, 7 and 15 days after operation, the cognitive function of the TIVA group was better than that of the control group, with statistically significance (P < .05, respectively). On the day of surgery, the two groups had similar sleep quality. The sleep quality of the TIVA group was better than that of the control group on 1, 3 and 7 days after surgery (P < .05, respectively). On the day of surgery, the levels of melatonin, cortisol, S-100ß protein and IL-6 in the two groups were equivalent (P > .05). On 1, 3, 7 and 15 days after surgery, cortisol and IL-6 in the TIVA group were lower than those of the control group, and melatonin was higher than that of the control group (P < .05, respectively). On 1, 3 and 7 days after operation, the S-100 ß protein in the TIVA group was lower than that in the control group (P < .05, respectively). CONCLUSION: Propofol-based TIVA has little effect on the cognitive function and sleep quality of elderly patients after surgery, and it is worthy of clinical application.

PromPDD, a web-based tool for the prediction, deciphering and design of promiscuous peptides that bind to HLA class I molecules.

Promiscuous peptides that can be presented by multiple human leukocyte antigens (HLAs) have great potential for the development of vaccines with wide population coverage. However, the current available methods for the prediction of peptides that bind to major histocompatibility complex (MHC) are mainly aimed at the rapid or mass screening of potential T cell epitopes from pathogen antigens or proteomics. The current approaches do not allow deciphering the contribution of the residue at each peptide position to the promiscuous binding ability of the peptide or obtaining guidelines for the design of promiscuous peptides. In this study, we re-evaluated and characterized four matrix-based prediction models that have been extensively used for the prediction of HLA-binding peptides and found that the prediction models generated based on the average relative binding (ARB) matrix shared a consistent and conservative threshold for all well-studied HLA class I alleles. Evaluations performed using datasets of HLA supertype-specific peptides with various cross-binding abilities and peptide mutant analogues indicated that the ARB-based binding matrices could be used to decipher and design promiscuous peptides that bind to multiple HLA molecules. A web-based tool called PromPDD was developed using ARB matrix-based models, and this tool enables the prediction, deciphering and design of promiscuous peptides that bind to multiple HLA molecules within or across HLA supertypes in a simpler and more direct manner. Furthermore, we expanded the application of PromPDD to HLA class I alleles with limited experimentally verified data by generating pan-specific matrices using a derived modular method, and 2641 HLA molecules encoded by HLA-A and HLA-B genes are available in PromPDD. PromPDD, which is freely available at http://www.immunoinformatics.net/PromPDD/, is the first tool for the deciphering and design of promiscuous peptides that bind to HLA class I molecules.

Should oral anticoagulant therapy be continued during dental extraction? A meta-analysis.

BACKGROUND: Oral anticoagulation therapy is widely used to reduce the risks of thromboembolism. However, the therapy increases the risk of hemorrhage during the surgical procedures. The aim of this meta-analysis was to evaluate the bleeding risk of patients continuing or discontinuing oral anticoagulant therapy while undergoing dental extractions. METHODS: Six electronic databases, including PubMed, Embase, Cochrane library, Web of Science, China Biology Medicine disc (CBM), and China National Knowledge Infrastructure (CNKI), were searched in March, 2016. Relevant articles were screened by two independent reviewers under our inclusion criteria. Quality was evaluated using the Cochrane Collaboration risk of bias tool. Meta-analyses were conducted with fixed and random effects models as appropriate. RESULTS: Six studies (with a total of 591 patients) were included in our meta-analysis. Our results showed that there was no significant difference in the bleeding risk between patients continuing or discontinuing oral anticoagulant therapy while undergoing dental extractions (risk ratio, 1.31; 95 % CI, 0.79, 2.14; P > 0.05). There was also no significant difference in bleeding risk 1 day (risk ratio, 0.91; 95 % CI, 0.35, 2.37; P > 0.05) and 7 days (risk ratio, 1.47; 95 % CI, 0.83, 2.59; P > 0.05) after the dental extraction. CONCLUSION: Under current studies and evidence, it appears that patients continuing oral anticoagulant therapy do not have an increased risk of bleeding after dental extractions compared to patients who discontinue oral anticoagulant therapy.

Association of TCR-signaling pathway with the development of lacrimal gland benign lymphoepithelial lesions.

AIM: To identify the association of the T cell receptor (TCR) signaling with the development of benign lymphoepithelial lesions (BLEL) of the lacrimal gland. METHODS: We collected affected lacrimal gland tissues from 9 patients who underwent dacryoadenectomy in the Capital Medical University Beijing Tongren Hospital Eye Center between August 2010 and March 2013 and were confirmed to have lacrimal gland BLEL by histopathological analysis. Tumor tissues from 9 patients with orbital cavernous hemangioma were also collected and used as control. Whole genome gene expression microarray was used to compare gene expression profiles of affected lacrimal gland tissues from patients with lacrimal gland BLEL to those from of orbital cavernous hemangiomas. Differential expression of TCR pathway genes between these tissues was confirmed by polymerase chain reaction (PCR) and immunohistochemistry. RESULTS: Microarray analysis showed that in lacrimal glands with BLEL, 32 signaling pathways were enriched in the upregulated genes, while 25 signaling pathways were enriched in the downregulated genes. In-depth analysis of the microarray data showed that the expression of 27 genes of the TCR signaling pathway increased significantly. To verify the differential expression of three of these genes, CD3, CD4, and interleukin (IL)-10, reverse transcription-PCR (RT-PCR) and immunohistochemistry assays were performed. RT-PCR analysis showed that CD3 and CD4 were expressed in the lacrimal glands with BLEL, but IL-10 was not expressed. Immunohistochemistry confirmed that CD3 and CD4 proteins were also present, but IL-10 protein was not. CD3, CD4, or IL-10 expression was not found in the orbital cavernous hemangiomas with either RT-PCR or immunohistochemistry. CONCLUSION: TCR signaling pathway might be involved in the pathogenesis of lacrimal gland BLEL.

Clinical research on benign lymphoepithelial lesions of lacrimal gland in 20 Chinese patients.

BACKGROUND: Benign lymphoepithelial lesion (BLEL) is characterized by symmetric bilateral swelling of the lacrimal and salivary glands and considered a subtype of immunoglobulin G4 (IgG4)-related sclerosing disease, the etiology and pathogenesis of which has not been determined. The purpose of the present study was to analyze the clinical features of BLEL in the lacrimal gland and the relationship between the serum level of IgG4 and BLEL. METHODS: Twenty consecutive patients with BLEL diagnosed in Department of Ophthalmology at Beijing Tongren Hospital, Capital Medical University between January 2012 and December 2013 were observed. The clinical features, imaging findings, laboratory tests, treatments and follow-up status of these 20 consecutive patients were analyzed. RESULTS: In all 20 patients, the ratio of male to female was 1:4, the ages ranged from 28 to 57 years, the ratio of unilateral to bilateral eyes involvement was 1:4, and painless uncongested symmetrical swelling of the upper eyelid was the main clinical manifestation. Orbital magnetic resonance imaging (MRI) showed that all patients involved lacrimal gland, which were obviously enlarged with equal signals in T1W images and T2W images and obvious enhancement on contrast MRI. Extraocular muscles were involved in 5 patients, salivary gland in 8 patients, and frontal nerve in 3 patients. Serum IgG4 concentration was elevated in 18 patients. The treatment strategy mainly included surgery and steroid administration. Three patients were lost to follow-up, 17 patients reached complete response, and no recurrence was observed. CONCLUSIONS: Eyelid swelling is the typical symptom of BLEL. Most of the patients involved bilateral lacrimal glands. High serum IgG4 level and abundant IgG4-positive plasma cell infiltration are the important features, which can be found in most of BLEL patients. Surgery combined with glucocorticoids is an efficient treatment strategy.

Facile preparation of gold nanocages and hollow gold nanospheres via solvent thermal treatment and their surface plasmon resonance and photothermal properties.

Although template etching method is one of the most common ways of preparation of hollow gold nanostructures, this approach still requires further improvements to avoid the collapse of gold shells after the cores were removed. In this work, an improved template etching method, with which hollow gold nanostructure is fabricated by etching Polystyrene (PS) cores from PS@Au core-shell nanospheres with solvent thermal treatment in N,N-Dimethylformamide (DMF), is demonstrated. When PS cores were removed by a thermal treatment process, gold nanoshells reconstruct and the collapse of the nanoshells is avoided. Gold nanocages and hollow gold nanospheres are easily obtained from the various structures of PS@Au core-shell nanospheres. These hollow nanostructures represent special near infrared (NIR) optical property and photothermal property. Compared with hollow gold nanospheres, the gold nanocages show higher temperature increase at the same particle concentration.

Shape separation of gold nanoparticles using a pH-responsive amphiphilic dendrimer according to their shape anisotropy distinction.

We present a novel method for shape-selective separation of nanoparticles in aqueous media utilizing an amphiphilic cationic dendrimer (C18N5) through regulation of the pH of the dispersion medium. This amphiphilic dendrimer (C18N5) bears a poly(amidoamine) dendron with four terminal amine groups and one hydrophobic long alkyl chain, and it can strongly adsorb to the surface of gold nanoparticles at low pH and promote their aggregation at high pH. During aggregation, isotropic NPs with smaller inter-particle contact area have much better solution stability than anisotropic NPs. Therefore, isotropic NPs remain stable in the supernatant, while anisotropic NPs are subject to aggregation and subsequent precipitation. As a result of this simple separation strategy, gold nanoplates with a purity level of 97% (in number density) can be rapidly and conveniently achieved.

Kimura's disease of the lacrimal gland mimicking IgG4-related orbital disease.

BACKGROUND: Kimura's disease (KD) is a rare and benign chronic inflammatory soft tissue disorder of unknown origin, which predominantly inflicts young male adults in Asia. IgG4-related disease is a new disease concept, established this century and characterized by fibrosis and sclerosis of the involved organs, with infiltration of IgG4-positive plasma cells. These two kinds of diseases share similar characteristics, which may complicate their diagnosis. CASE PRESENTATION: A 47-year-old Chinese man presented to our Department of Ophthalmology with a 26-month history of painless swelling and redness left upper eyelid. Surgical excisions of the left lacrimal gland were performed. A histopathology examination showed follicular hyperplasia with reactive germinal centres and eosinophilic infiltration involving the interfollicular areas as well as proliferation of post capillary venules, all signs of Kimura disease. Immunohistochemical analysis of the cells demonstrated positive staining for CK, Vimentin, CD3, CD4, CD20, CD21, CD117, CD5, CD8, CD23, IgG and IgG4 (30 per high-power field) and negative staining for CD10 and CD34. Some ophthalmologists in our department questioned whether the histological and immunohistochemical findings were also compatible with features of IgG4-related diseases. There was no sign of recurrence during the twelve months of regular follow-up. CONCLUSION: Kimura's disease may present with high serum IgG4 levels, which may be an epiphenomenon related to chronic antigen exposure. As clinical doctors, especially ophthalmologists, we should recognize the possibility of the occurrence of increased serum levels of IgG4 in Kimura's disease to ensure correct diagnosis.

Photothermal therapy of cancer cells using novel hollow gold nanoflowers.

This article presents a new strategy for fabricating large gold nanoflowers (AuNFs) that exhibit high biological safety under visible light and very strong photothermal cytotoxicity to HeLa cells under irradiation with near-infrared (NIR) light. This particular type of AuNF was constructed using vesicles produced from a multiamine head surfactant as a template followed by depositing gold nanoparticles (AuNPs) and growing their crystallites on the surface of vesicles. The localized surface plasmon-resonance spectrum of this type of AuNF can be easily modulated to the NIR region by controlling the size of the AuNFs. When the size of the AuNFs increased, biosafety under visible light improved and cytotoxicity increased under NIR irradiation. Experiments in vitro with HeLa cells and in vivo with small mice have been carried out, with promising results. The mechanism for this phenomenon is based on the hypothesis that it is difficult for larger AuNFs to enter the cell without NIR irradiation, but they enter the cell easily at the higher temperatures caused by NIR irradiation. We believe that these effects will exist in other types of noble metallic NPs and cancer cells. In addition, the affinity between AuNPs and functional biomolecules, such as aptamers and biomarkers, will make this type of AuNF a good recognition device in cancer diagnosis and therapy.

Synthesis of highly branched gold nanodendrites with a narrow size distribution and tunable NIR and SERS using a multiamine surfactant.

Gold nanodendrites with a long and densely branched morphology were fabricated by a seed-mediated method in a solution containing gold nanoparticles (AuNPs), bis(amidoethyl-carbamoylethyl)octadecylamine (C18N3), HAuCl4, and the reducing agent ascorbic acid (AA). The length and density of the branches could be mediated by changing the AuNP seed and AA concentrations. The amphiphilic C18N3 molecules function as a template and induce the unique morphology of the AuNPs/C18N3 structures. The localized surface plasmon resonance (LSPR) peaks of the gold nanodendrites can be modulated from the visible (∼530 nm) to the near-infrared region (∼1100 nm) of the electromagnetic spectrum. Surface-enhanced Raman scattering (SERS) signals using rhodamine can also be mediated by changing the seed and AA concentrations. These unique highly branched gold nanodendrites with a narrow size distribution and tunable NIR and SERS spectra should have great potential in sensing applications.

A new strategy for the controlled deposition of gold nanoparticle aggregates on two-dimensional polystyrene arrays and its application in glucose oxidase immobilization.

Nano/microstructures play an important role in nanoparticle applications. This paper describes an innovative strategy to fabricate a variety of gold nanoparticle aggregates (AuNPs) on large-scale arrays of up to ∼1 cm(2) made from polystyrene (PS). A dendritic surfactant, C18N3, has multi-amine head groups that can control the thickness of a double layer adsorbed on the PS sphere surface in a pH-dependent manner. Controlling the pH and immersion time in the C18N3 solution allows the morphology of AuNPs deposited on the PS spheres (PS@AuNP) to be regulated. The influence of nano/microstructures on the activity enhancement of glucose oxidase (GOD) was investigated. The results indicated that well-ordered PS@AuNP arrays performed much better in the specific activity enhancement of GOD compared with free GOD and GOD immobilized on PS arrays. Furthermore, it was observed that the immobilized GOD on 2D PS@AuNP arrays maintained a highly improved operational stability compared to free GOD. The mechanism behind this effect is discussed. For practical applications, prepared PS@AuNP arrays can be used as an effective chip for GOD immobilization and application.

Effects of aggregation and the surface properties of gold nanoparticles on cytotoxicity and cell growth.

The effect of gold nanoparticles (Au NPs) on cells remains open for investigation. Here we show that small Au NPs can be endocytosed by cells and form aggregates inside the cell, resulting in cytotoxicity. When the aggregates become too large to enter the cell and instead adhere onto the cell surface, however, the growth rate of HeLa cells increases. Printed patterns of Au NPs fabricated through inkjet printing technology were used to study the effects of Au NP aggregation on human cervical carcinoma (HeLa) cell activity. The growth of the HeLa cells was inhibited on the polymer-coated Au NPs but increased on the silicon substrate. On the uncoated Au NP surface, however, the HeLa cell growth rate was higher than that on the silicon substrate. Experiments with Escherichia coli cells showed a similar effect of the Au NPs. This phenomenon provides a new perspective for research on toxicity in nanoparticle biology. FROM THE CLINICAL EDITOR: Printed patterns of Au NPs fabricated through inkjet printing technology were used to study the effects of Au NP aggregation on human cervical carcinoma (HeLa) cell activity. Small Au NPs can be endocytosed by cells resulting in cytotoxicity; in contrast, large aggregates adhere onto the cell surface and increase the growth rate of HeLa cells.

Cognitive impairment is common in Parkinson's disease without dementia in the early and middle stages in a Han Chinese cohort.

BACKGROUND: Cognitive impairments have been reported to be common in Parkinson's disease (PD) without dementia, which occur not only in the late stages of PD, but also in the early and middle stages. Until now, no reports on the profile of cognitive impairment in Chinese non-demented PD population have been published yet. Different ethnic groups should be assessed to improve evaluation of cognitive impairment in clinical practice. The aims of this study are to estimate the frequencies and profile of cognitive impairments and to explore the risk factors of cognitive impairments in Han Chinese non-demented PD patients at early and middle stages. METHODS: Eighty non-demented PD patients in early and middle stages and 86 healthy controls were invited to participate in this study. Neuropsychological batteries testing executive function, visuospatial function, memory and attention were evaluated. Cognitive impairments were defined as impaired performance in at least one cognitive domain. RESULTS: Neuropsychological batteries detected 30 cases with executive dysfunction, 27 cases with memory impairment, eight cases with visuospatial dysfunction and seven cases with attention impairment. As many as 48 cases (60%) of PD patients presented cognitive impairment. Logistic regression analysis indicated that education level and Hoehn & Yahr stage were associated with cognitive impairment in PD. CONCLUSIONS: Cognitive impairment is common in the early and middle stages of PD without dementia; executive function is the most common domain impaired in a Chinese PD population. Cognitive impairment might be predicted by lower education level and higher Hoehn and Yahr stage.

Epigenetic involvement of Alien/ESET complex in thyroid hormone-mediated repression of E2F1 gene expression and cell proliferation.

The ligand-bound thyroid hormone receptor (TR) is known to repress via a negative TRE (nTRE) the expression of E2F1, a key transcription factor that controls the G1/S phase transition. Alien has been identified as a novel interacting factor of E2F1 and acts as a corepressor of E2F1. The detailed molecular mechanism by which Alien inhibits E2F1 gene expression remains unclear. Here, we report that the histone H3 lysine 9 (H3K9) methyltransferase (HMT) ESET is an integral component of the corepressor Alien complex and the Alien/ESET complex is recruited to both sites, the E2F1 and the nTRE site of the E2F1 gene while the recruitment to the negative thyroid hormone response element (nTRE) is induced by the ligand-bound TRß1 within the E2F1 gene promoter. We show that, overexpression of ESET promotes, whereas knockdown of ESET releases, the inhibition of TRß1-regulated gene transcription upon T3 stimulation; and H3K9 methylation is required for TRß1-repressed transcription. Furthermore, depletion of ESET impairs thyroid hormone-repressed proliferation as well as the G1/S transition of the cell cycle. Taken together, our data indicate that ESET is involved in TRß1-mediated transcription repression and provide a molecular basis of thyroid hormone-induced repression of proliferation.

[Significant improvement of motor symptoms by deep brain stimulation of bilateral subthalamic nucleus in patients with moderate or advanced Parkinson's disease].

OBJECTIVE: To study the effects of deep brain stimulation (DBS) of bilateral subthalamic nucleus (STN) on the motor and non-motor symptoms in moderate or advanced Parkinson's disease (PD) patients. METHODS: From August 2006 to January 2010, 21 consecutive PD patients with refractory motor fluctuations or dyskinesia underwent operations at our hospital. All patients were evaluated by unified Parkinson's disease rating scale (UPDRS), Hoehn & Yahr (H&Y) stage, Parkinson's disease questionnaire (PDQ-39), mini mental state examination (MMSE), Parkinson's disease sleep scale-Chinese vision (PDSS-CV), Pittsburgh sleep quality index (PSQI), Hamilton depression rating scale (HAMD) and Hamilton anxiety rating scale (HAMA). And the daily dosage of dopaminergic agents was recorded at 1 week pre-operation and 3, 6 and 12 months post-operation. RESULTS: Ten patients finished a 12-month follow-up. Their motor functions showed significant improvement. And the scores of UPDRS-motor, tremor, rigidity, bradykinesia and axial symptoms reduced significantly in the on-stimulation-off-medication condition and the on-stimulation-on-medication condition vs the on-medication condition pre-operation. And the improvement of tremor was the most pronounced (52.1% and 77.7% respectively). The H&Y stage decreased significantly from 3.2 ± 0.7 to 2.5 ± 0.4 post-operation. The activities of daily living improved while PDQ-39 declined significantly from 56 ± 9 pre-operation to 32 ± 13 at 12 months follow-up. The score changes of MMSE, PDSS-CV, PSQI, HAMA and HAMD were statistically insignificant. The levo-dopa equivalent dose of 1-year post-operation decreased significantly by 49.2% versus that of pre-operation (P < 0.05). CONCLUSION: Bilateral STN-DBS can significant ameliorate the motor symptoms of moderate or advanced PD patients, reduce the dosage of anti-PD medications and improve the quality of life. This procedure has the advantages of a greater safety, minor side effects and an easy controllability.

Contraction-related stimuli regulate GLUT4 traffic in C2C12-GLUT4myc skeletal muscle cells.

Muscle contraction stimulates glucose uptake acutely to increase energy supply, but suitable cellular models that faithfully reproduce this complex phenomenon are lacking. To this end, we have developed a cellular model of contracting C(2)C(12) myotubes overexpressing GLUT4 with an exofacial myc-epitope tag (GLUT4myc) and explored stimulation of GLUT4 traffic by physiologically relevant agents. Carbachol (an acetylcholine receptor agonist) induced a gain in cell surface GLUT4myc that was mediated by nicotinic acetylcholine receptors. Carbachol also activated AMPK, and this response was sensitive to the contractile myosin ATPase inhibitor N-benzyl-p-toluenesulfonamide. The gain in surface GLUT4myc elicited by carbachol or by the AMPK activator 5-amino-4-carboxamide-1 beta-ribose was sensitive to chemical inhibition of AMPK activity by compound C and partially reduced by siRNA-mediated knockdown of AMPK catalytic subunits or LKB1. In addition, the carbachol-induced gain in cell surface GLUT4myc was partially sensitive to chelation of intracellular calcium with BAPTA-AM. However, the carbachol-induced gain in cell surface GLUT4myc was not sensitive to the CaMKK inhibitor STO-609 despite expression of both isoforms of this enzyme and a rise in cytosolic calcium by carbachol. Therefore, separate AMPK- and calcium-dependent signals contribute to mobilizing GLUT4 in response to carbachol, providing an in vitro cell model that recapitulates the two major signals whereby acute contraction regulates glucose uptake in skeletal muscle. This system will be ideal to further analyze the underlying molecular events of contraction-regulated GLUT4 traffic.