Impact of uranium on antibiotic resistance in activated sludge.

The emergence of antibiotic resistance genes (ARGs) and antibiotic-resistant bacteria (ARB) in the environment is well established as a human health crisis. The impact of radioactive heavy metals on ecosystems and ultimately on human health has become a global issue, especially for the regions suffering various nuclear activities or accidents. However, whether the radionuclides can affect the fate of antibiotic resistance in bacteria remains poorly understood. Here, the dynamics of ARB, three forms of ARGs-intracellular ARGs (iARGs), adsorbed extracellular ARGs (aeARGs), and free extracellular ARGs (feARGs)-and microbial communities were investigated following exposure to uranium (U), a representative radioactive heavy metal. The results showed that 90-d of U exposure at environmentally relevant concentrations of 0.05 mg/L or 5 mg/L significantly increased the ARB concentration in activated sludge (p < 0.05). Furthermore, 90-d of U exposure slightly elevated the absolute abundance of aeARGs (except tetO) and sulfonamide iARGs, but decreased tetracycline iARGs. Regarding feARGs, the abundance of tetC, tetO, and sul1 decreased after 90-d of U stress, whereas sul2 showed the opposite trend. Partial least-squares path model analysis revealed that the abundance of aeARGs and iARGs under U stress was predominantly driven by increased cell membrane permeability/intI1 abundance and cell membrane permeability/reactive oxygen species concentration, respectively. Conversely, the changes in feARGs abundance depended on the composition of the microbial community and the expression of efflux pumps. Our findings shed light on the variations of ARGs and ARB in activated sludge under U exposure, providing a more comprehensive understanding of antibiotic resistance risks aggravated by radioactive heavy metal-containing wastewater.

Multi-joint protective effects of lumbar brace on lumbar, hip, knee, and ankle in parachute landing with backpack load.

There were high injury risks on lumbar and lower limb joints in parachuting landing, and the lumbar brace could protect lumbar. Besides, a backpack load was necessary in parachute landing and increased the injury risk. This study aimed to evaluate multi-joints protective effects of the lumbar brace on lumbar and lower limb joints in parachuting landing with the backpack load. Seven participants landed from a 120 cm height platform without and with a lumbar brace and without and with a 5-kg backpack load, respectively. Infrared makers were pasted on trunk, pelvis, and lower limb in order to build a multi-rigid-body model for calculating kinematic and kinetic parameters. The joint angular displacements of lumbar and ankle and the peak vertical ground reaction force were significantly decreased from 29.2 ± 9.2°, 45.2 ± 7.8°, and 14.7 ± 2.0 bodyweight to 21.6 ± 4.9° (p < 0.05), 39.0 ± 10.1° (p < 0.05) and 10.7 ± 1.3 bodyweight (p < 0.05) respectively by the lumbar brace with no backpack load, and the joint angular displacement of hip was significantly increased from 52.6 ± 7.2° to 68.3 ± 12.5° (p < 0.05). The joint angular displacement of lumbar and ankle were significantly decreased from 29.0 ± 5.0° and 53.8 ± 5.1° to 25.1 ± 5.2° (p < 0.05) and 48.5 ± 2.5° (p < 0.05) respectively by the lumbar brace with the backpack load, and the joint angular displacement of hip and knee were significantly increased from 60.1 ± 8.2° and 110.1 ± 9.3° to 69.7 ± 13.2° (p < 0.05) and 116.8 ± 5.8° (p < 0.05), respectively. The lumbar brace could provide the multi-joint protective effect by decreasing injuries of lumbar and ankle in landing both without and with the backpack load.

Finite element analysis of shank and ankle with different boot collar heights in parachuting landing on inversion ground surface.

This study aimed to the biomechanics of the foot-ankle-shank complex with different boots collar heights in parachuting landing on inversion ground surface. A finite element model including tibia, fibula, ankle, foot and parachuting boot was developed. Three collar heights (low, medium, high) of the parachuting boot were simulated. Von-Mises stress, ankle inversion angle, ligament force and bone displacement were analyzed. Compared with that of the high and low collar heights, boots with medium collar height produced the lowest peak stress on the tibia and the articular cartilage of the subtalar joint. In addition, the medium collar height can better control the ankle inversion and minimize the tensile forces on the lateral ankle ligaments.

Protection by Ankle Brace for Lower-Extremity Joints in Half-Squat Parachuting Landing With a Backpack.

Half-squat parachuting landing is a kind of activity with high impact force. Injuries on lower-extremity joints are common in half-squat parachuting landing and would be increased with a backpack. An ankle brace was used to prevent ankle injuries in landing. However, few quantitative studies reported about the protection of an ankle brace for lower-extremity joints in half-squat parachuting landing with a backpack. This study focused on evaluating the protective effects of an ankle brace in half-squat parachuting landing with a backpack. Seven male participants landed from 120 cm with a backpack and an ankle brace. Each participant performed three landing trials on every experimental condition. Kinetics and kinematics of the hip, knee, and ankle were analyzed. It was found that the ankle brace did not significantly affect the ground reaction force with backpack but increased the ground reaction force from 14.7 ± 2.0 bodyweight to 16.2 ± 1.9 bodyweight (p = 0.017) without the backpack. The ankle brace significantly (p < 0.05) decreased the angular displacement, angular velocity, and angular acceleration of the ankle both without and with the backpack. In conclusion, the ankle brace could restrict ankle motion and significantly increase ground reaction force without the backpack. However, the ankle brace did not significantly influence ground reaction force and still restricted ankle motion with the backpack. Therefore, the ankle brace was more effective in half-squat parachuting landing with the backpack than no-backpack landing.

Finite element analysis of lumbar spine with different backpack positions in parachuting landing.

The purpose of this study was to investigate the lumbar spine stress with different backpack positions in parachuting landing using a finite element model of lumbar vertebra 1-5. The backpack gravity center was set at three positions (posterior-high (case PH), posterior-low (case PL), and anterior-low (case AL)) respectively. In results, the peak Von-Mises stresses of the matrix, nucleus, fibers, endplate and ligament in case AL were 2.765 MPa, 0.534 MPa, 6.561 MPa, 4.045 MPa and 1.790 MPa respectively, lower than those in case PL (6.913 MPa, 1.316 MPa, 20.716 MPa, 10.917 MPa and 5.147 MPa respectively) and case PH (7.328 MPa, 1.394 MPa, 22.147 MPa, 11.617 MPa and 5.464 MPa respectively). In conclusion, setting the gravity center of backpack at anterior-low position would reduce lumbar spine stress and reduce lumbar spine injuries.

Erratum to "Analysis of Bone Mineral Density/Content of Paratroopers and Hoopsters".

[This corrects the article DOI: 10.1155/2018/6030624.].

Kinematics and kinetics of lower-extremity joints in parachuting landing with backpack and knee brace.

Injuries on lower-extremity joints were caused by high impact force in parachuting landing. Knee brace was used to protect knee by restraining motion of knee. Backpack was necessary in parachuting landing and would increase lower-extremity joints injuries. This study aimed to analyze kinematics and kinetics of hip, knee and ankle for investigating multi-joint protection of knee brace for those joints in parachuting landing with backpack. Seven participants landed from 120 cm height. Kinematics and kinetics of hip, knee and ankle were analyzed. It was found that without backpack knee brace decreased angular displacements of hip (12.0%), knee (10.3%) and ankle (18.6%) on sagittal plane and angular velocities of hip (11.9%), knee (6.6%) and ankle (20.9%) on sagittal plane. With backpack, knee brace decreased angular displacement (5.5%) and angular velocity of knee (6.2%) on sagittal plane, but did not significantly influence those of hip and ankle on sagittal plane. Ground reaction force, joint moments and joint energy absorptions were not significantly influenced with knee brace. In conclusion, in parachuting landing without backpack, knee brace could provide multi-joint protection for hip, knee and ankle. In parachuting landing with backpack, knee brace could still protect knee, but could not protect hip and ankle.

Analysis of Bone Mineral Density/Content of Paratroopers and Hoopsters.

The different mechanical stimulus affects the bone mass and bone strength. The aim of this study was to investigate the effect of landing posture of the hoopster and paratrooper on the bone mass. In this study, 39 male participants were recruited including 13 paratroopers, 13 hoopsters, and 13 common students (control groups). Bone area (BA), BMD and BMC of calcaneus, and 1-5th of the metatarsus, hip, and lumbar spine (L1-L4) were measured by the dual-energy X-ray absorptiometry. Also, the vertical ground reaction forces (GRFs) of hoopsters and paratroopers were measured by the landing of 1.2 m 3D force platform. BA of hoopsters at the calcaneus, lumbar spine, and hip were significantly higher than the control group. The lumbar spine, hip, calcaneus, the 1st and 2nd metatarsals, BMC of paratroopers, and control groups were significantly lower than hoopsters. BMD of the lumbar spine, hip, and right and left femoral necks in hoopsters were significantly higher than the other participants. BMC and BMD of lower limber showed no significant difference between paratroopers and the control group. Besides, peak GRFs of paratroopers (11.06 times of BW) were significantly higher than hoopsters (6.49 times of BW). The higher GRF in the landing train is not always in accordance with higher BMD and BMC. Variable loads in hoopsters can improve bone remodeling and play an important role in bone expansions for trabecular bones. This will be considered by the method of training to prevent bone loss.

Strain measurement of objects subjected to aerodynamic heating using digital image correlation: experimental design and preliminary results.

In thermomechanical testing of hypersonic materials and structures, direct observation and quantitative strain measurement of the front surface of a test specimen directly exposed to severe aerodynamic heating has been considered as a very challenging task. In this work, a novel quartz infrared heating device with an observation window is designed to reproduce the transient thermal environment experienced by hypersonic vehicles. The specially designed experimental system allows the capture of test article's surface images at various temperatures using an optical system outfitted with a bandpass filter. The captured images are post-processed by digital image correlation to extract full-field thermal deformation. To verify the viability and accuracy of the established system, thermal strains of a chromiumnickel austenite stainless steel sample heated from room temperature up to 600 °C were determined. The preliminary results indicate that the air disturbance between the camera and the specimen due to heat haze induces apparent distortions in the recorded images and large errors in the measured strains, but the average values of the measured strains are accurate enough. Limitations and further improvements of the proposed technique are discussed.

Synergistic action of a RAF inhibitor and a dual PI3K/mTOR inhibitor in thyroid cancer.

PURPOSE: In thyroid cancer clinical trials, agents targeting VEGF receptors (VEGFR) and RET, among other kinases, have led to partial responses but few complete or durable responses. The RAF-MEK-ERK and PI3K-AKT-mTOR signaling pathways are frequently activated in differentiated and medullary thyroid cancer (DTC and MTC) and may provide therapeutic targets for these diseases. We tested a novel drug combination targeting RAF, phosphoinositide 3-kinase (PI3K), and mTOR, plus VEGFR2 and RET, in thyroid cancer preclinical models with defined genetic backgrounds. EXPERIMENTAL DESIGN: RAF265, an ATP-competitive pan-RAF inhibitor active against VEGFR2, and BEZ-235, a PI3K inhibitor also active against Torc1 and Torc2, were tested alone and in combination in a panel of thyroid cancer lines. We tested RAF265 and BEZ-235 for kinase inhibition, growth inhibition and cell-cycle alterations, and inhibition of signaling targets and tumor growth in xenograft models. RESULTS: Both drugs potently inhibited their kinase targets in the extracellular signal-regulated kinase (ERK) and PI3K pathways. In addition, RAF265 had significant RET inhibitory activity (IC50 = 25-50 nmol/L for RET(C634W)). The combination strongly inhibited proliferation of DTC and MTC cell lines with mutations in RAS, BRAF, PTEN, and RET. Synergy was shown for B-CPAP (BRAF(V600E)) and TT cells (RET(C634W)). The combination of both drugs significantly inhibited growth of CAL62 (KRAS(G12R/G12R)) and TT xenografts, thoroughly inhibiting ERK and PI3K pathway signaling. CONCLUSIONS: Combined blockade of ERK and PI3K signaling potently inhibits growth in preclinical models representing the key genotypes seen in refractory thyroid cancer. These targets and therapies are promising for further development in both differentiated and medullary thyroid cancers.

Dual inhibition of mitogen-activated protein kinase kinase and mammalian target of rapamycin in differentiated and anaplastic thyroid cancer.

CONTEXT: Differentiated thyroid cancer and anaplastic thyroid cancer tumors frequently have activation of the ras/raf /MAPK kinase (MEK)/ERK and phosphatidylinositol 3-kinase (PI-3K)/AKT/mammalian target of rapamycin (mTOR) signaling pathways. OBJECTIVE: The objective of the study was to investigate the efficacy of MEK and mTOR inhibitors in preclinical thyroid cancer treatment models with defined mutation status. EXPERIMENTAL DESIGN: The MEK inhibitor AZD6244 (ARRY-142886) and mTOR inhibitor rapamycin were tested separately and in combination in 10 differentiated thyroid cancer and anaplastic thyroid cancer cell lines and in a xenograft model for evidence of pathway inhibition, growth inhibition, apoptosis, and long-range adaptation and resistance. RESULTS: Seven of 10 tested lines had evidence of significant basal activity of the PI-3K/AKT/mTOR pathway, with elevated phosphorylated AKT and phosphorylated p70 S6 kinase. Activation of ras/RAF/MEK/ERK was equally common in this panel. All 10 lines exhibited better than 60% growth inhibition with combined MEK and mTOR inhibition, including lines with BRAF, Ret-PTC, ras, and PTEN mutations. Rapamycin or AZD6244 alone achieved this threshold in six and two lines, respectively. Dual-pathway inhibition in the Ret-PTC mutant cell line TPC1 caused an intense G(1) arrest in cell culture and reversible cytostatic inhibition in a xenograft model. We did not observe significant feedback up-regulation of AKT activation in either acute or prolonged exposures. CONCLUSION: These preclinical results support the inclusion of thyroid cancer patients in early-phase clinical trials combining RAS/RAF/MEK/ERK and PI-3K/AKT/mTOR pathway inhibition.

Achaete-scute complex homologue 1 regulates tumor-initiating capacity in human small cell lung cancer.

The basic helix-loop-helix transcription factor achaete-scute complex homologue 1 (ASCL1) is essential for the development of normal lung neuroendocrine cells as well as other endocrine and neural tissues. Small cell lung cancer (SCLC) and non-SCLC with neuroendocrine features express ASCL1, where the factor may play a role in the virulence and primitive neuroendocrine phenotype of these tumors. In this study, RNA interference knockdown of ASCL1 in cultured SCLC resulted in inhibition of soft agar clonogenic capacity and induction of apoptosis. cDNA microarray analyses bolstered by expression studies, flow cytometry, and chromatin immunoprecipitation identified two candidate stem cell marker genes, CD133 and aldehyde dehydrogenase 1A1 (ALDH1A1), to be directly regulated by ASCL1 in SCLC. In SCLC direct xenograft tumors, we detected a relatively abundant CD133(high)-ASCL1(high)-ALDH1(high) subpopulation with markedly enhanced tumorigenicity compared with cells with weak CD133 expression. Tumorigenicity in the CD133(high) subpopulation depended on continued ASCL1 expression. Whereas CD133(high) cells readily reconstituted the range of CD133 expression seen in the original xenograft tumor, CD133(low) cells could not. Our findings suggest that a broad range of SCLC cells has tumorigenic capacity rather than a small discrete population. Intrinsic tumor cell heterogeneity, including variation in key regulatory factors such as ASCL1, can modulate tumorigenicity in SCLC.

Regulation of androgen receptor activity by tyrosine phosphorylation.

The androgen receptor (AR) is essential for the growth of prostate cancer cells. Here, we report that tyrosine phosphorylation of AR is induced by growth factors and elevated in hormone-refractory prostate tumors. Mutation of the major tyrosine phosphorylation site in AR significantly inhibits the growth of prostate cancer cells under androgen-depleted conditions. The Src tyrosine kinase appears to be responsible for phosphorylating AR, and there is a positive correlation of AR tyrosine phosphorylation with Src tyrosine kinase activity in human prostate tumors. Our data collectively suggest that growth factors and their downstream tyrosine kinases, which are elevated during hormone-ablation therapy, can induce tyrosine phosphorylation of AR and such modification may be important for prostate tumor growth under androgen-depleted conditions.

Bi-directional regulation between tyrosine kinase Etk/BMX and tumor suppressor p53 in response to DNA damage.

Etk/Bmx, a member of the Tec family of nonreceptor tyrosine kinases, has been implicated in the regulation of various cellular processes including proliferation, differentiation, motility, and apoptosis. Here, we report the identification of Tec family kinases as the potential interacting proteins of the tumor suppressor p53 by an Src homology 3 domain array screening. Etk is physically associated with p53 through its Src homology 3 domain and the proline-rich domain of p53. Induction of p53 expression by DNA damage inhibits Etk activity in several cell types. Down-regulation of Etk expression by a specific small interfering RNA sensitizes prostate cancer cells to doxorubicin-induced apoptosis, suggesting that inhibition of Etk activity is required for apoptosis in response to DNA damage. We also show that Etk primarily interacts with p53 in the cytoplasm and that such interaction leads to bidirectional inhibition of the activities of both proteins. Overexpression of Etk in prostate cancer cells results in inhibition of p53 transcriptional activity and its interaction with the mitochondrial protein BAK and confers the resistance to doxorubicin. Therefore, we propose that the stoichiometry between p53 and the Tec family kinases in a given cell type may determine its sensitivity to chemotherapeutic drugs.

Synergism of cytoplasmic kinases in IL6-induced ligand-independent activation of androgen receptor in prostate cancer cells.

IL6 is a pleiotropic cytokine which has been implicated in ligand-independent activation of androgen receptor in prostate cancer cells. Here, we present the evidence that two cytoplasmic kinases Pim1 and Etk are involved in this process. We showed that Pim1 is expressed in all prostate cancer cell lines examined. Both the expression level and the kinase activity of Pim1 are regulated by IL6 in these cells. Furthermore, we showed that IL6 downstream tyrosine kinase Etk can induce tyrosine phosphorylation of Pim1 which is correlated with its kinase activity. Mutation of the conserved Tyrosine 218 in the activation loop results in reduced kinase activity of Pim1. Interestingly, Etk can also be activated by Pim1 when they are coexpressed in prostate cancer cells, suggesting a possible positive feedback loop between Etk and Pim1. It appears that both Pim1 and Etk are required for IL6-induced activation of androgen receptor-mediated transcription in prostate cancer cells because overexpression of the kinase-deficient form of either Pim1 or Etk dramatically blocks the IL6 effect. Coexpression of the two kinases together but neither one alone is sufficient to activate ARE-containing promoter. Taken together, our data suggest a synergism of Ser/Thr kinase Pim1 and tyrosine kinase Etk in IL6 signaling and provide new insights into ligand-independent activation of androgen receptor in prostate cancer cells.

Interaction between Src and a C-terminal proline-rich motif of Akt is required for Akt activation.

Activation of Akt by growth factors is a multistep process. Here, we provide evidence that tyrosine kinase Src is directly associated with Akt through the interaction between its SH3 domain and a conserved proline-rich motif (PXXP) in the C-terminal regulatory region of Akt. Substitution of the proline residues Pro-424 and Pro-427 by alanines results in loss of Akt activity and phosphorylation induced by the epidermal growth factor (EGF), possibly because these mutations disrupt the interaction between Akt and the SH3 domain of Src. This possibility is corroborated by our observation that the Akt mutant lacking these two prolines fails to bind to Src both in vivo and in vitro. We also showed that phosphorylation of Tyr-315 in Akt induced by Src or EGF is dependent on the integrity of this proline-rich motif. Furthermore, the Akt mutant lacking this proline motif fails to block the transcription activity of Forkhead in 293 cells and poorly stimulates the proliferation of Madin-Darby canine kidney cells. Taken together, our data suggest that the interaction between the SH3 domain of Src family kinases and the proline-rich motif in the C-terminal regulatory region of Akt is required for tyrosine phosphorylation of Akt and its subsequent activation. It is noteworthy that this PXXP motif is conserved throughout several members of AGC kinase family, implying that association of this motif with the SH3 domain of an upstream regulator may represent a general mechanism applicable to these kinases as well.

Effects of testosterone and 17-beta-estradiol on TNF-alpha-induced E-selectin and VCAM-1 expression in endothelial cells. Analysis of the underlying receptor pathways.

This study investigated the effects of testosterone and 17-beta-estradiol on tumor necrosis factor-alpha (TNF-alpha)-induced endothelial expression of E-selectin and vascular cell adhesion molecule-1 (VCAM-1) and the potential roles of hormone receptors involved in these actions. Human umbilical vein endothelial cells (HUVEC) were stimulated with TNF-alpha in the presence or absence of testosterone or 17-beta-estradiol, and the expression of E-selectin and VCAM-1 was investigated. As shown by Western blot analysis, co-administration with testosterone or 17-beta-estradiol increased the expression of E-selectin and VCAM-1 induced by TNF-alpha at 6 h and 3 h, respectively. Similarly, RT-PCR analysis revealed a significant increase in the amount of mRNA for E-selectin and VCAM-1 after co-administration with testosterone or 17-beta-estradiol in TNF-alpha-stimulated HUVEC. The presence of mRNA and proteins for androgen receptor and estrogen receptor alpha in HUVEC was verified by RT-PCR and Western blot. Flow cytometric analysis showed that preincubation with androgen receptor antagonist cyproterone and estrogen receptor antagonist tamoxifen completely abrogated the upregulating effects of testosterone and 17-beta-estradiol on TNF-alpha-induced E-selectin and VCAM-1 expression, respectively. Expression of TNF receptors in TNF-alpha-stimulated HUVEC was not influenced by testosterone and 17-beta-estradiol. The data indicate that both testosterone and 17-beta-estradiol increase TNF-alpha-induced E-selectin and VCAM-1 expression in endothelial cells via a receptor-mediated system, and expression of TNF receptors are not changed in these actions. The implications of these results for the facilitory effects of both sex hormones on immune reactions are discussed.