Composite bone graft of CaO-MgO-SiO2 glass-ceramics and CaSO4 ceramics for boosting bone formation rate.

This study develops a composite bone graft of CaO-MgO-SiO2 glass-ceramic and CaSO4 [abbreviated as (CMS)3-x(CS)x] via the sponge replication technique with weight fractions of x = 0, 1, 1.5, 2, and 3. The (CMS)1.5(CS)1.5 composite displays a superior degradability and, a suitable compressive strength of ∼3 MPa, and excellent cell proliferation and differentiation. The in vivo rat femur test in the hybrid-pore (CMS)1.5(CS)1.5 composite granules achieves a higher rate of bone formation, which is ∼2.7 times better than that of the commercial HAP/ß-TCP at 12 weeks. Improved expressions of osteocyte and mature osteocyte marker genes, namely (Spp1, Dmp1, and Fgf23), were observed in the (CMS)1.5(CS)1.5 group, indicating a faster differentiation into mature bone tissue. The ions release of (CMS)1.5(CS)1.5 through the ERK1/2 signaling pathway promotes osteogenic differentiation. The high bone generation rate can be attributed to faster active ions release and modified surface topography. This work highlights an excellent bone graft candidate for clinical applications in orthopedic surgery.

Local application of zoledronate inhibits early bone resorption and promotes bone formation.

Nonunion resulting from early bone resorption is common after bone transplantation surgery. In these patients, instability or osteoporosis causes hyperactive catabolism relative to anabolism, leading to graft resorption instead of fusion. Systemic zoledronate administration inhibits osteoclastogenesis and is widely used to prevent osteoporosis; however, evidence on local zoledronate application is controversial due to osteoblast cytotoxicity, uncontrolled dosing regimens, and local release methods. We investigated the effects of zolendronate on osteoclastogenesis and osteogenesis and explored the corresponding signaling pathways. In vitro cytotoxicity and differentiation of MC3T3E1 cells, rat bone marrow stromal cells (BMSCs) and preosteoclasts (RAW264.7 cells) were evaluated with different zolendronate concentrations. In vivo bone regeneration ability was tested by transplanting different concentrations of zolendronate with ß-tricalcium phosphate (TCP) bone substitute into rat femoral critical-sized bone defects. In vitro, zolendronate concentrations below 2.5 × 10-7 M did not compromise viability in the three cell lines and did not promote osteogenic differentiation in MC3T3E1 cells and BMSCs. In RAW264.7 cells, zoledronate inhibited extracellular regulated protein kinases and c-Jun n-terminal kinase signaling, downregulating c-Fos and NFATc1 expression, with reduced expression of fusion-related dendritic cell­specific transmembrane protein and osteoclast-specific Ctsk and tartrate-resistant acid phosphatase (. In vivo, histological staining revealed increased osteoid formation and neovascularization and reduced fibrotic tissue with 500 µM and 2000 µM zolendronate. More osteoclasts were found in the normal saline group after 6 weeks, and sequential osteoclast formation occurred after zoledronate treatment, indicating inhibition of bone resorption during early callus formation without inhibition of late-stage bone remodeling. In vivo, soaking ß-TCP artificial bone with 500 µM or 2000 µM zoledronate is a promising approach for bone regeneration, with potential applications in bone transplantation.

Meteorology-driven PM2.5 interannual variability over East Asia.

Atmospheric fine particulate matter (PM2.5) is a human health risk factor, but its ambient concentration depends on both precursor emissions and meteorology. While emission reductions are used to set PM2.5-related health policies, the effect of meteorology is often overlooked. To explore this aspect, we examined PM2.5 interannual variability (IAV) associated with meteorological parameters using the long-term simulation from the Community Earth System Model (CESM1), a global climate-chemistry model, with fixed emissions. The results are subsequently contrasted with the MERRA-2 reanalysis dataset, which inherently considers emission and meteorology effects. Over continental East Asia, the CESM1 domain-average PM2.5 IAV is 6.7 %, mainly attributed to humidity, precipitation, and ventilation variation. The grid-cell PM2.5 IAVs over southern East China are larger, up to 12 % due to the more substantial influence of El Niño-induced meteorological anomalies. Under such climate extreme, sub-regional PM2.5 concentration may occasionally exceed WHO air quality guideline levels despite the compliance of the long-term mean. The simulated PM2.5 IAV over continental East Asia is ~25 % of that derived from the MERRA-2 data, which highlights the influence of both emission and meteorology-driven variations and trends inherent in the latter. Although emission-driven variability is significant to PM2.5 IAV, in remote areas downwind of major source regions in East Asia, North America, and Western Europe, the MERRA-2 data revealed that meteorological variations contributed more to PM2.5 IAV than emission variations. Thus, when setting policies for complying with the WHO PM2.5-related air quality guideline levels, the highest annual PM2.5 associated with climate extremes should be considered instead of that based on average climate conditions.

Prevalence and Clinical Implications of Mismatch Repair-Proficient Colorectal Cancer in Patients With Lynch Syndrome.

PURPOSE: Lynch syndrome (LS)-associated colorectal cancer (CRC) is characterized by mismatch repair-deficiency (MMR-D) and/or microsatellite instability (MSI). However, with increasing utilization of germline testing, MMR-proficient (MMR-P) and/or microsatellite stable (MSS) CRC has also been observed. We sought to characterize MMR-P/MSS CRC among patients with LS. METHODS: Patients with solid tumors with germline MMR pathogenic/likely pathogenic (P/LP) variants were identified on a prospective matched tumor-normal next-generation sequencing (NGS) protocol. CRCs were evaluated for MMR-D via immunohistochemical (IHC) staining and/or MSI via NGS. Clinical variables were correlated with MMR status using nonparametric tests. RESULTS: Among 17,617 patients with solid tumors, 1.4% (n = 242) had LS. A total of 36% (86 of 242) of patients with LS had at least one CRC that underwent NGS profiling, amounting to 99 pooled CRCs assessed. A total of 10% (10 of 99) of CRCs were MMR-P, with 100% concordance between MSS status and retained MMR protein staining. A total of 89% (8 of 9) of patients in the MMR-P group had MSH6 or PMS2 variants, compared with 30% (23 of 77) in the MMR-D group (P = .001). A total of 46% (6 of 13) of PMS2+ patients had MMR-P CRC. The median age of onset was 58 and 43 years for MMR-P and MMR-D CRC, respectively (P = .07). Despite the later median age of onset, 40% (4 of 10) of MMR-P CRCs were diagnosed <50. A total of 60% (6 of 10) of MMR-P CRCs were metastatic compared with 13% (12 of 89) of MMR-D CRCs (P = .002). A total of 33% (3 of 9) of patients with MMR-P CRC did not meet LS testing criteria. CONCLUSION: Patients with LS remained at risk for MMR-P CRC, which was more prevalent among patients with MSH6 and PMS2 variants. MMR-P CRC was later onset and more commonly metastatic compared with MMR-D CRC. Confirmation of tumor MMR/MSI status is critical for patient management and familial risk estimation.

A Rare PDGFRA Exon 15 Germline Mutation Identified in a Patient With Phenotypic Manifestations Concerning for GIST-Plus Syndrome: A Case Report and Review of Literature.

Molecular alterations in PDGFRA are well-described as drivers of sporadic gastrointestinal stromal tumors (GISTs) and inflammatory fibroid polyps (IFPs). However, a small number of families with germline PDGFRA mutations in exons 12, 14, and 18 have been reported, forming the basis of an autosomal dominant inherited disorder with incomplete penetrance and variable expressivity, now referred to as PDGFRA-mutant syndrome or GIST-plus syndrome. Phenotypic manifestations of this rare syndrome include multiple gastrointestinal GISTS, IFPs, fibrous tumors, and other variable features. Herein, we report the case of a 58-year-old female who presented with a gastric GIST and numerous small intestinal IFPs, found to harbor a previously undescribed germline PDGFRA exon 15 p.G680R mutation. Somatic tumor testing was performed on the GIST, a duodenal IFP, and an ileal IFP utilizing a targeted next-generation sequencing panel, revealing additional and distinct secondary PDGFRA exon 12 somatic mutations in each of the 3 tumors. Our findings raise important considerations regarding mechanisms of tumor development in patients with underlying germline PDGFRA alterations and highlight the potential utility of expanding currently available germline and somatic testing panels to include exons outside the typical hotspot regions.

The role of counter-torque holders in tightening of pedicle screw-rod constructs: a biomechanical study in a porcine model.

BACKGROUND CONTEXT: Pedicle screw-rod assembly procedures following pedicle screw insertion include contouring and placing rods into screw tulips, introducing set screws into the tulip along the screw thread, applying a counter-torque holder and tightening all the set screws clockwise. Even if an appropriate pedicle screw is implanted, screw dislodgement after tightening of the tulip and set screw is not uncommon. Pedicle wall violation resulting from excessive rotational force due to inadequate use of a counter-torque holder might be the reason. However, the strain change in the pedicle during tulip-set screw tightening and the role of counter-torque have never been investigated. PURPOSE: This study determined differences in the strain change in the outer and inner pedicle walls during tulip-set screw tightening; additionally, the influence of counter-torque on pedicle wall violation was elucidated. STUDY DESIGN: A controlled biomechanical study; the strain values of outer and inner pedicle walls in cadaveric porcine L4-L5 vertebrae during tulip-set screw tightening with or without a counter-torque holder were measured. METHODS: Twelve L4-L5 fresh-frozen porcine lumbar vertebrae were implanted with screw-rod constructs; the set screw was randomly locked into the tulip in the right L5, right L4, left L5 and left L4 testing groups. The maximal values from eight strain gauges (P-R-O: outer cortex of right pedicle in proximal vertebra; P-R-I: inner cortex of right pedicle in proximal vertebra; D-R-O: outer cortex of right pedicle in distal vertebra; D-R-I: inner cortex of right pedicle in distal vertebra; P-L-O: outer cortex of left pedicle in proximal vertebra; P-L-I: inner cortex of left pedicle in proximal vertebra; D-L-O: outer cortex of left pedicle in distal vertebra; D-L-I: outer cortex of left pedicle in proximal vertebra) for each specimen during tightening to 12 Nm were measured. RESULTS: The maximal strain values of the ipsilateral strain gauges in all testing groups were almost significantly higher when a counter-torque holder was not used than when one was used. The strain values in the adjacent pedicle of specimens without a counter-torque holder were significantly increased: P-R-O and P-R-I in the right L5 group; D-R-I in the right L4 group; P-L-I and P-L-O in the left L5 group; D-L-O and D-L-I in the left L4 group. CONCLUSIONS: The constraint effect of counter-torque during tulip-set screw tightening is necessary. Clockwise rotational force with a fragile lateral pedicle wall suggests that caution is required when using a counter-torque holder to tighten the right L5 and left L4 constructs. CLINICAL SIGNIFICANCE: A counter-torque holder is important during tulip-set screw tightening; improper use may lead to adjacent pedicle wall violation, sequentially resulting in pedicle screw loosening.

Detecting mismatch repair deficiency in solid neoplasms: immunohistochemistry, microsatellite instability, or both?

In managing patients with solid tumors, the value of detecting the status of tumor DNA mismatch repair function is widely recognized. Mismatch repair protein immunohistochemistry and molecular microsatellite instability testing constitute the two major test modalities currently in use, yet each is associated with caveats and limitations that can be consequential. Most notably, the traditional approach of defining mismatch repair protein immunohistochemistry abnormality by complete loss of staining in all tumor cells is evolving. Partial or clonal loss is becoming recognized as a manifestation of gene abnormality; in some cases, such clonal loss is associated with germline pathogenic variants. The current criteria and cutoff values for defining microsatellite instability-high are developed primarily according to colorectal tumors. Non-colorectal cases, and occasionally even colorectal tumors, that are mismatch repair-deficient by immunohistochemistry but not microsatellite instability-high by current standards are being recognized. Emerging data suggest that these immunohistochemistry abnormal / non-microsatellite instability-high cases warrant further genetic workup for Lynch syndrome detection. Whether these tumors respond to immunotherapy is a question still to be addressed. It is imperative that pathologists as well as clinicians and investigators be aware of such intricacies regarding routine immunohistochemistry and microsatellite instability testing and the results they generate. This review summarizes our current understanding of the advantages and limitations of these tests and offer our view on what constitutes the most optimal strategy in test selection and how best to utilize case context to enhance the interpretation of the test results.

Paediatric non-alcoholic fatty liver disease: an approach to pathological evaluation.

Non-alcoholic fatty liver disease (NAFLD) is becoming an increasingly important healthcare issue along with the rising rates of obesity worldwide. It is the most common chronic liver disease in the paediatric population and the fastest growing indication for liver transplant in young adults. The pathogenesis is complex with contributions from multiple factors and genetic predisposition. While non-invasive laboratory tests and imaging modalities are being increasingly used, the liver biopsy continues to play a crucial role in the diagnosis and prognosis of NAFLD. Histologically, the assessment of paediatric fatty liver disease requires special considerations with respect to a periportal predominant pattern seen in prepubertal patients, as well as a different set of disease processes in the differential diagnosis. In this review, we provide a summary of current knowledge on the epidemiology, pathogenesis and clinical course of paediatric NAFLD as well as the clinical guidelines on diagnosis and management. We discuss the indications and limitations of liver biopsy, histological patterns seen in paediatric NAFLD, other entities to be considered in the differential diagnosis, and conclude with appropriate triaging of liver biopsies and essential elements of pathology reporting.

Electrospinning of Quaternized Chitosan-Poly(vinyl alcohol) Composite Nanofiber Membrane: Processing Optimization and Antibacterial Efficacy.

N-(2-hydroxy) propyl-3-trimethylammonium chitosan chloride (HTCC) is a type of quaternary ammonium chitosan derivative with an antibacterial activity superior to the pristine chitosan, but its electrospinnability is limited. In this study, polyvinyl alcohol (PVA) was blended with HTCC to improve the electrospinnability of nanofibers. The electrospinning of PVA-HTCC nanofiber membranes was optimized in terms of structural stability and antimicrobial performance. Based on scanning electron microscopic analysis, the morphology and diameter of the produced nanofibers were influenced by the applied voltage, flow rate of the feed solution, and weight ratio of the polymer blend. An increase in the HTCC content decreased the average nanofiber diameter. The maximum water solubility of the PVA-HTCC nanofibers reached the maximum value of 70.92% at 12 h and 25 °C. The antibacterial activity of PVA-HTCC nanofiber membranes against Escherichia coli was ~90%, which is significantly higher than that of PVA-chitosan nanofiber membrane. Moreover, the antibacterial efficiency of PVA-HTCC nanofiber membranes remained unaffected after 5 cycles of antibacterial treatment. The good antibacterial performance and biocompatibility of PVA-HTCC nanofiber membrane makes them attractive for biomedical and biochemical applications that necessitate sterile conditions.

From 3D printing to 3D bioprinting: the material properties of polymeric material and its derived bioink for achieving tissue specific architectures.

The application of 3D printing technologies fields for biological tissues, organs, and cells in the context of medical and biotechnology applications requires a significant amount of innovation in a narrow printability range. 3D bioprinting is one such way of addressing critical design challenges in tissue engineering. In a more general sense, 3D printing has become essential in customized implant designing, faithful reproduction of microenvironmental niches, sustainable development of implants, in the capacity to address issues of effective cellular integration, and long-term stability of the cellular constructs in tissue engineering. This review covers various aspects of 3D bioprinting, describes the current state-of-the-art solutions for all aforementioned critical issues, and includes various illustrative representations of technologies supporting the development of phases of 3D bioprinting. It also demonstrates several bio-inks and their properties crucial for being used for 3D printing applications. The review focus on bringing together different examples and current trends in tissue engineering applications, including bone, cartilage, muscles, neuron, skin, esophagus, trachea, tympanic membrane, cornea, blood vessel, immune system, and tumor models utilizing 3D printing technology and to provide an outlook of the future potentials and barriers.

Cytotoxicity and cell response of preosteoblast in calcium sulfate-augmented PMMA bone cement.

Poly(methyl methacrylate) (PMMA) has been widely used in orthopedic applications, but bone ingrowth and toxic monomer release are drawback of this material. Particle reinforcement with osteoconductive substitute, such as calcium sulfate (CaSO4), is one of the solutions used to modify PMMA bone cement. The current study investigated the mechanical, chemical and biological properties of CaSO4-augmented bone cement. Mechanical strength was measured by a material testing machine. The concentration of methyl methacrylate (MMA) monomer from the various formulations of PMMA mixed with CaSO4was measured by ultra-performance liquid chromatography (UPLC). CCK-8 assay and ALP assay were performed to evaluate cytotoxicity of released MMA monomer and cell differentiation. The attachment of cells to CaSO4-augmented bone cement discs was observed by confocal and scanning electron microscopy, and surface topography was also evaluated by atomic force microscopy. The results revealed that increased CaSO4weight ratios led to compromised mechanical strength and increased MMA monomer release. Cell density and cell differentiation on CaSO4-augmented bone cement discs were decreased at CaSO4weight ratios above 10%. In addition, the presence of micropores on the surface and surface roughness were both increased for PMMA composite discs containing higher levels of CaSO4. These results demonstrated that fewer MC3T3-E1 cells on the surface of CaSO4-PMMA composites was correlated to increased MMA monomer release, micropore number and surface roughness. In summary, the augmentation of a higher proportion of CaSO4(>10 wt. %) to PMMA did not promote the biological properties of traditional PMMA bone cement.

Enhanced mechanical and biological performances of CaO-MgO-SiO2 glass-ceramics via the modulation of glass and ceramic phases.

This work reports a new CaO-MgO-SiO2 (CMS) bioactive glass-ceramic, using ZrO2 as a nucleus to modulate the ratios of glass and ceramic phases as a function of sintering temperature. Mg-rich bioactive CMS glass-ceramics exhibit advantages regarding mechanical strength (flexural strength ~190 MPa and compressive strength ~555 MPa), in-vitro and in-vivo biocompatibilities, and bone ingrowth. The high mechanical strengths could be attributed to the CaMgSi2O6 glass-ceramic and lower porosity. X-ray absorption spectra indicate an increased SiO covalent bond via the development of CaMgSi2O6 glass-ceramics. From the in-vitro cytotoxicity and BMSC differentiation assays, the CMS samples sintered above 800 °C exhibited better cell attachment and differentiation, possibly due to structural stability, appropriate pore, and ion release to boost osteogenesis. Compared to hydroxyapatite (HA) ceramics, the CMS glass-ceramics display higher mechanical strengths, biocompatibility, and osteoconductivity. An in-vivo experiment demonstrated a fine bone-ingrowth profile around the CMS implant. This study may further the application of CMS glass-ceramics in bone implants.

Antibacterial efficacy of quaternized chitosan/poly (vinyl alcohol) nanofiber membrane crosslinked with blocked diisocyanate.

N-[(2-hydroxyl-3-trimethylammonium) propyl] chitosan chloride (HTCC), which is a type of chitosan derivative with quaternary ammonium groups, possesses a higher antibacterial activity as compared to the pristine chitosan. The nanofiber membranes made of HTCC are attractive for applications demanding for antibacterial function. However, the hydrophilic nature of HTCC makes it unsuitable for electrospinning of nanofibers. Hence, biodegradable polyvinyl alcohol (PVA) was proposed as an additive to improve the electrospinnability of HTCC. In this work, PVA/HTCC nanofiber membrane was crosslinked with the blocked diisocyanate (BI) to enhance the stability of nanofiber membrane in water. Microbiological assessments showed that the PVA/HTCC/BI nanofiber membranes possessed a good antibacterial efficacy (∼100 %) against E. coli. Moreover, the biocompatibility of PVA/HTCC/BI nanofiber membrane was proven by the cytotoxicity test on mouse fibroblasts. These promising results indicated that the PVA/HTCC/BI nanofiber membrane can be a promising material for food packaging and as a potential wound dressing for skin regeneration.

Antibacterial efficacy of poly(hexamethylene biguanide) immobilized on chitosan/dye-modified nanofiber membranes.

This study aimed to develop a novel electrospun polyacrylonitrile (PAN) nanofiber membrane with the enhanced antibacterial property. The PAN nanofiber membrane was first subjected to alkaline hydrolysis treatment, and the treated membrane was subsequently grafted with chitosan (CS) to obtain a CS-modified nanofiber membrane (P-COOH-CS). The modified membrane was then coupled with different dye molecules to form P-COOH-CS-Dye membranes. Lastly, poly(hexamethylene biguanide) hydrochloride (PHMB) was immobilized on the modified membrane to produce P-COOH-CS-Dye-PHMB. Physical characterization studies were conducted on all the synthesized nanofiber membranes. The antibacterial efficacies of nanofiber membranes prepared under different synthesis conditions were evaluated systematically. Under the optimum synthesis conditions, P-COOH-CS-Dye-PHMB was highly effective in disinfecting a high concentration of Escherichia coli, with an antibacterial efficacy of approximately 100%. Additionally, the P-COOH-CS-Dye-PHMB exhibited an outstanding wash durability as its antibacterial efficacy was only reduced in the range of 5%-7% even after 5 repeated cycles of treatment. Overall, the experimental results of this study suggested that the P-COOH-CS-Dye-PHMB is a promising antibacterial nanofiber membrane that can be adopted in the food, pharmaceutical, and textile industries.

Phosphorylation of PDHA by AMPK Drives TCA Cycle to Promote Cancer Metastasis.

Cancer metastasis accounts for the major cause of cancer-related deaths. How disseminated cancer cells cope with hostile microenvironments in secondary site for full-blown metastasis is largely unknown. Here, we show that AMPK (AMP-activated protein kinase), activated in mouse metastasis models, drives pyruvate dehydrogenase complex (PDHc) activation to maintain TCA cycle (tricarboxylic acid cycle) and promotes cancer metastasis by adapting cancer cells to metabolic and oxidative stresses. This AMPK-PDHc axis is activated in advanced breast cancer and predicts poor metastasis-free survival. Mechanistically, AMPK localizes in the mitochondrial matrix and phosphorylates the catalytic alpha subunit of PDHc (PDHA) on two residues S295 and S314, which activates the enzymatic activity of PDHc and alleviates an inhibitory phosphorylation by PDHKs, respectively. Importantly, these phosphorylation events mediate PDHc function in cancer metastasis. Our study reveals that AMPK-mediated PDHA phosphorylation drives PDHc activation and TCA cycle to empower cancer cells adaptation to metastatic microenvironments for metastasis.

Cell migration of preosteoblast cells on a clinical gelatin sponge for 3D bone tissue engineering.

Using three-dimensional (3D) bone engineering to fabricate bone segments is a better choice for repairing bone defects than using autologous bone. However, biomaterials for bone engineering are burdened with some clinical safety concerns. In this study, we layered commonly found clinical materials, hemostatic gelatin sponges, in a novel manner to create a 3D scaffold for bone engineering purposes. We further examined the comparable benefits of our design with both closed- and open-bottom holders. Cells in stacked layer disc systems were examined after a week of growth and differentiation. Osteoblasts in the outer layers of both closed- and open-bottom holder systems displayed gradually increased alkaline phosphatase (ALP) activity but decreased osteopontin (OPN) expression. Further, cell proliferation assays and LIVE/DEAD staining revealed decreased viable cell counts in the top layer with increased incubation time. However, while layered disc systems with closed-bottom holders underwent differentiation, they kept more differentiated cells alive within the gelatin sponge disc scaffold after 28 d of culturing. Whether cells were inoculated into the top, middle, or bottom portions of the layered disc stack, osteoblasts showed a preference for migrating to the top layer, in keeping with the oxygen and nutrients gradients. Regarding practical application, this study offers valuable information to promote the use of hemostatic gelatin sponges for bone engineering.

Incorporation of surface-modified hydroxyapatite into poly(methyl methacrylate) to improve biological activity and bone ingrowth.

Poly(methyl methacrylate) (PMMA) is the most frequently used bone void filler in orthopedic surgery. However, the interface between the PMMA-based cement and adjacent bone tissue is typically weak as PMMA bone cement is inherently bioinert and not ideal for bone ingrowth. The present study aims to improve the affinity between the polymer and ceramic interphases. By surface modifying nano-sized hydroxyapatite (nHAP) with ethylene glycol and poly(ɛ-caprolactone) (PCL) sequentially via a two-step ring opening reaction, affinity was improved between the polymer and ceramic interphases of PCL-grafted ethylene glycol-HAP (gHAP) in PMMA. Due to better affinity, the compressive strength of gHAP/PMMA was significantly enhanced compared with nHAP/PMMA. Furthermore, PMMA with 20 wt.% gHAP promoted pre-osteoblast cell proliferation in vitro and showed the best osteogenic activity between the composites tested in vivo. Taken together, gHAP/PMMA not only improves the interfacial adhesion between the nanoparticles and cement, but also increases the biological activity and affinity between the osteoblast cells and PMMA composite cement. These results show that gHAP and its use in polymer/bioceramic composite has great potential to improve the functionality of PMMA cement.

The critical role of AMPK in driving Akt activation under stress, tumorigenesis and drug resistance.

PI3K/Akt signaling is activated in cancers and governs tumor initiation and progression, but how Akt is activated under diverse stresses is poorly understood. Here we identify AMPK as an essential regulator for Akt activation by various stresses. Surprisingly, AMPK is also activated by growth factor EGF through Ca2+/Calmodulin-dependent kinase and is essential for EGF-mediated Akt activation and biological functions. AMPK phosphorylates Skp2 at S256 and promotes the integrity and E3 ligase activity of Skp2 SCF complex leading to K63-linked ubiquitination and activation of Akt and subsequent oncogenic processes. Importantly, AMPK-mediated Skp2 S256 phosphorylation promotes breast cancer progression in mouse tumor models, correlates with Akt and AMPK activation in breast cancer patients, and predicts poor survival outcomes. Finally, targeting AMPK-mediated Skp2 S256 phosphorylation sensitizes cells to anti-EGF receptor targeted therapy. Our study sheds light on how stress and EGF induce Akt activation and new mechanisms for AMPK-mediated oncogenesis and drug resistance.