Author Correction: mTORC2-AKT signaling to ATP-citrate lyase drives brown adipogenesis and de novo lipogenesis.

An amendment to this paper has been published and can be accessed via a link at the top of the paper.

mTORC2-AKT signaling to ATP-citrate lyase drives brown adipogenesis and de novo lipogenesis.

mTORC2 phosphorylates AKT in a hydrophobic motif site that is a biomarker of insulin sensitivity. In brown adipocytes, mTORC2 regulates glucose and lipid metabolism, however the mechanism has been unclear because downstream AKT signaling appears unaffected by mTORC2 loss. Here, by applying immunoblotting, targeted phosphoproteomics and metabolite profiling, we identify ATP-citrate lyase (ACLY) as a distinctly mTORC2-sensitive AKT substrate in brown preadipocytes. mTORC2 appears dispensable for most other AKT actions examined, indicating a previously unappreciated selectivity in mTORC2-AKT signaling. Rescue experiments suggest brown preadipocytes require the mTORC2/AKT/ACLY pathway to induce PPAR-gamma and establish the epigenetic landscape during differentiation. Evidence in mature brown adipocytes also suggests mTORC2 acts through ACLY to increase carbohydrate response element binding protein (ChREBP) activity, histone acetylation, and gluco-lipogenic gene expression. Substrate utilization studies additionally implicate mTORC2 in promoting acetyl-CoA synthesis from acetate through acetyl-CoA synthetase 2 (ACSS2). These data suggest that a principal mTORC2 action is controlling nuclear-cytoplasmic acetyl-CoA synthesis.

Synthesis of isobemisiose, neosartose, and fischerose: three α-1,6-linked trehalose-based oligosaccharides identified from Neosartorya fischeri.

Three complex α-1,6-linked trehalose-based oligosaccharides with unique preservation properties, isobemisiose, neosartose, and fischerose, were recently identified from the extreme stress-tolerant ascospores of Neosartorya fischeri. Herein, we report the first concise, scalable, and iterative chemical synthesis of these oligosaccharides from a differentially protected thioglycoside donor and a selectively protected, asymmetric trehalose acceptor. This work constitutes an improved synthesis of isobemisiose, and is also the first reported synthesis of neosartose, a tetrasaccharide, and fischerose, a pentasaccharide, in good yield. Importantly, in-depth studies of biological function are enabled by this synthetic platform.

Regulation of lipid synthesis by the RNA helicase Mov10 controls Wnt5a production.

Expression of the Wnt ligand Wnt5a is frequently elevated in melanoma and is thought to be a critical regulator of cell movement during metastasis. However, the mechanisms regulating its expression are unknown. We find that the level of secreted Wnt5a varies by as much as 10-fold between cell lines and correlates more strongly with invasion than total cellular levels. Our results indicate that the RNA helicase Mov10 plays a role in Wnt5a synthesis and secretion. Inhibition of Mov10 increases secreted Wnt5a levels in melanoma cells by increasing Wnt5a synthesis and acylation. This is achieved by increasing fatty acid synthase (FASN) and stearoyl-CoA desaturase expression, leading to elevated levels of palmitoleoyl-CoA, required for Wnt ligand lipid modification and secretion. Melanoma tumors exhibit reduced expression of Mov10 compared with benign nevi and Mov10 levels inversely correlate with FASN levels in primary tumors. These results reveal a previously unappreciated role for aberrant lipid metabolism in regulating Wnt5a signaling that may be a critical step in melanoma progression.

Harmful algal bloom toxins alter c-Fos protein expression in the brain of killifish, Fundulus heteroclitus.

The immediate early gene c-fos, and its protein product c-Fos, are known to be induced in neurons of mammals and fish as a result of neuronal stimulation. The purpose of this study was to quantitatively examine CNS alterations in killifish, Fundulus heteroclitus, in relation to harmful algal bloom (HAB) toxin exposure. c-Fos expression was visualized using immunocytochemistry in the brains of killifish exposed to the excitatory neurotoxins domoic acid (DA) and brevetoxin (PbTx-2), and a paralytic neurotoxin, saxitoxin (STX), released from HABs. In addition, a simulated transport stress experiment was conducted to investigate effects of physical stress on c-Fos induction. Groups of fish were exposed to the different stress agents, brain sections were processed for c-Fos staining, and expression was quantified by brain region. Fish exposed to DA, STX, and transport stress displayed significant alterations in neuronal c-Fos expression when compared to control fish (p< or = 0.05). DA, PbTx-2, and transport stress increased c-Fos expression in the optic tecta regions of the brain, whereas STX significantly decreased expression. This is the first study to quantify c-Fos protein expression in fish exposed to HAB toxins. General alterations in brain activity, as well as knowledge of specific regions within the brain activated in association with HABs or other stressors, provides valuable insights into the neural control of fish behavior as well as sublethal effects of specific stressors in the CNS.

Trigger finger release with hand surface landmark ratios: an anatomic and clinical study.

The purpose of this study was to identify surface landmark ratios to locate the A1 pulley and clarify the controversy of differing anatomic descriptions of the A1, C0, and A2 pulleys. Minimally invasive and percutaneous approaches to A1 pulley release may be facilitated with surface landmark ratios, which identify and predict the proximal and distal margins of the A1 pulley. Two-hundred fifty-sixty fingers were dissected in 64 preserved cadaver hands. Measurements of A1 pulley lengths and pulley margins in relation to surface landmarks were obtained. We found that the distance from the palmar digital crease to the proximal interphalangeal crease (mean, 2.42 +/- 0.03 cm) corresponds to the distance of the proximal edge of the A1 pulley from the palmar digital crease (mean, 2.45 +/- 0.03 cm). The mean absolute difference between these two measured distances in each finger was 0.13 cm, with a 95 percent confidence interval of 0.11 to 0.14 cm. Thus, the distance between the palmar digital crease and the proximal interphalangeal crease can be used to predict the distance between the palmar digital crease and the A1 pulley proximal edge with reasonable accuracy. A1 pulley length averaged 0.98 +/- 0.02 cm for the small finger and 1.17 +/- 0.02 cm for the index, middle, and ring fingers. The length of the A1 pulley was significantly shorter (p < 0.001) for the small finger than for the index, middle, and ring fingers. Additionally, a cruciate (C0) pulley was consistently located between the A1 and A2 pulleys, an average of 0.46 cm proximal to the palmar digital crease, which can serve as guide for concluding the release of the A1 pulley. Clinically, hand surface landmark ratios were used to release 32 trigger fingers with a minimally invasive technique, without a complication during 4- to 30-week follow-up. We conclude that hand surface landmark ratios can serve to locate the proximal A1 pulley edge, thus facilitating complete trigger finger release by either open or minimally invasive techniques. Additionally, our study clarifies the discrepancy of prior smaller reports of the pulley system anatomy regarding the existence of the C0 pulley between the A1 and A2 pulleys. The cruciate fibers of this C0 pulley can serve as the distal boundary for release of trigger finger.

Bipedicle paraspinous muscle flaps for spinal wound closure: an anatomic and clinical study.

The purpose of this study was to evaluate the vascular anatomy of the paraspinous muscles and review their clinical use as bipedicled flaps in spinal wound closure. Anatomically, through cadaver dissections, lead oxide injections, and radiographic imaging, the blood supply to the paraspinous muscles was determined. Clinically, 29 consecutive patients treated with spinal wounds and exposed bone or hardware were reviewed retrospectively. Of these patients, 19 underwent closure in delayed primary fashion, whereas 10 were referred to plastic surgery for reconstruction because of the complex nature of their wounds. The cadaver study demonstrated the paraspinous muscles to possess a segmental arterial supply through medial and lateral perforators. Division of the medial perforators allowed for medial advancement of the muscles. Lead oxide injection of the lateral perforators demonstrated adequate medial muscle perfusion with ligation of the medial perforators. Ten of the 29 patients (six women, four men, 32 to 62 years of age) were reconstructed with paraspinous (eight), latissimus (one), and trapezius (one) muscle flaps. A higher complication rate was found in wounds closed in delayed primary fashion (13 of 19 patients, 68 percent) than those reconstructed with muscle flaps (2 of 10 patients, 20 percent) (p = 0.021). Follow-up of the muscle flap reconstructed patients averaged 12 months (range, 3 to 27 months). Cadaver muscle injections predicted and clinical cases confirmed that the paraspinous muscles can be raised on lateral perforators and advanced medially to close lumbar spine wounds reliably with fewer complications.

Five-year follow-up results of instep plantar fasciotomy for chronic heel pain.

The results of a 5-year follow-up of patients who underwent surgical partial release of plantar fascia with plantar skin incision for treatment of chronic heel pain are presented. Evaluation included survey results from both a modified Mayo Scoring System and a list of specific questions developed by the authors. Thirty-three feet of 30 patients are evaluated. Thirty of the 33 feet studied achieved good or excellent results, scoring 80 or better on a 100-point scale. Ninety percent pain relief was achieved in 27 of the 33 feet studied (81.8%). A satisfaction rate of 90% or better was found with 30 of the 33 feet studied (90.9%). Long-lasting complications were described by five patients, including opposite foot pain, dorsal foot pain after extended activity, scar tissue discomfort, callus at scar area, and continued heel pain.

The effect of inducible nitric oxide synthase (iNOS) inhibition on smoke inhalation injury in sheep.

Recent studies on smoke inhalation injury have been focused on nitric oxide (NO) as an essential factor of progressive lung injury. We studied the effects of inducible nitric oxide synthase (iNOS) inhibition on inhalation injury in sheep. Sheep (n = 14) were prepared surgically for chronic study. After recovery period, the sheep received 48 breaths of cotton smoke. The animals were then randomised into two groups: MEG group [30 mg/kg mercaptoethylguanidine (MEG), selective inhibitor of iNOS and peroxynitrite scavenger, was given 1 h after injury and then 8 h for 41 h, n = 7] and control group (0.9% NaCl, n = 7). All animals were ventilated mechanically, and airway blood flow was measured using colored microspheres. In the control group, following significant increase in airway blood flow, deterioration in the PaO2/FiO2 ratio was observed. Whereas in the MEG group, it was not observed. In addition, the MEG group did not show significant increase in pulmonary vascular resistance and intrapulmonary shunt fraction. Lung wet/dry ratios, a marker of pulmonary edema, were significantly lower in the MEG group. At 48 h after injury, lung tissue-conjugated dienes, an index of lung oxidative tissue injury, were significantly lower in the MEG group than in the control group. Our data suggest that 1) iNOS-NO produced in the airway circulation plays a major role on the significant increase in airway blood flow, which may contribute to the spread of injury from injured airway to the lung parenchyma; 2) iNOS-NO induced in the pulmonary circulation contributes to the loss of hypoxic pulmonary vasoconstriction; and 3) iNOS-NO plays an important role on the lung oxidative tissue injury.

Novel glycopeptide antibiotics: N-alkylated derivatives active against vancomycin-resistant enterococci.

LY264826 (A82846B) is a naturally-occurring glycopeptide antibiotic, differing from vancomycin in the stereochemistry of the amino-sugar of the disaccharide function, and the presence of a third sugar attached at the benzylic position of amino acid residue 6. Despite these seemingly subtle differences, LY264826 is approximately 10 times more active than vancomycin against the enterococci. In the pursuit of new antibiotics active against multiresistant Gram-positive organisms, an extensive side chain SAR was developed focusing on the reductive alkylation of LY264826 at the amino function of the disaccharide moiety. A new series of derivatives having varying degrees of structural diversity in the side chain (e.g. varying lengths and degrees of rigidity) was found to have potent activity against vancomycin-resistant enterococci (MIC's < 1.0 microgram/ml) as well as activity against staphylococci and streptococci as good or better than vancomycin.

Enzymatic deacylation of teicoplanin followed by reductive alkylation: synthesis and antibacterial activity of new glycopeptides.

Novel glycopeptides derived from teicoplanin were prepared and evaluated for activity against antibiotic-resistant gram-positive pathogens. Removal of the fatty acid sidechains of teicoplanin was accomplished by enzymatic deacylation. The resulting deacylated teicoplanin was subjected to reductive alkylation resulting in mono- and di-alkylated compounds at the 2 possible primary amines. Deacylated teicoplanin was less active than teicoplanin against enterococci and staphylococci (MIC > or =32 microg/ml). All mono- and di-alkylated products regained some activity, and some had potent activity against both staphylococci and glycopeptide-resistant enterococci. MICs of the most potent di-alkylated compounds ranged from 0.25 approximately 2 microg/ml against glycopeptide-resistant enterococci.

3-trifluoromethylcarbacephems: synthesis of broad spectrum antibacterial compounds.

The enhanced stability of the carbacephem nucleus over the corresponding cephalosporin nucleus has allowed the synthesis of 7-arylglycyl-3-trifluoromethyl-carbacephems. These unique carbacephems possess broad spectrum activity and high stability to both plasmid and chromosomally mediated beta-lactamases.

Structure-activity relationship of carbacephalosporins and cephalosporins: antibacterial activity and interaction with the intestinal proton-dependent dipeptide transport carrier of Caco-2 cells.

An intestinal proton-dependent peptide transporter located on the lumenal surface of the enterocyte is responsible for the uptake of many orally absorbed beta-lactam antibiotics. Both cephalexin and loracarbef are transported by this mechanism into the human intestinal Caco-2 cell line. Forty-seven analogs of the carbacephalosporin loracarbef and the cephalosporin cephalexin were prepared to evaluate the structural features necessary for uptake by this transport carrier. Compounds were evaluated for their antibacterial activities and for their ability to inhibit 1 mM cephalexin uptake and, subsequently, uptake into Caco-2 cells. Three clinically evaluated orally absorbed carbacephems were taken up by Caco-2 cells, consistent with their excellent bioavailability in humans. Although the carrier preferred the L stereoisomer, these compounds lacked antibacterial activity and were hydrolyzed intracellularly in Caco-2 cells. Compounds modified at the 3 position of cephalexin and loracarbef with a cyclopropyl or a trifluoromethyl group inhibited cephalexin uptake. Analogs with lipophilic groups on the primary amine of the side chain inhibited cephalexin uptake, retained activity against gram-positive bacteria but lost activity against gram-negative bacteria. Substitution of the phenylglycl side chain with phenylacetyl side chains gave similar results. Compounds which lacked an aromatic ring in the side chain inhibited cephalexin uptake but lost all antibacterial activity. Thus, the phenylglycl side chain is not absolutely required for uptake. Different structural features are required for antibacterial activity and for being a substrate of the transporter. Competition studies with cephalexin indicate that human intestinal Caco-2 cells may be a useful model system for initially guiding structure-activity relationships for the rational design of new oral agents.

Semisynthetic glycopeptide antibiotics derived from LY264826 active against vancomycin-resistant enterococci.

Certain derivatives of the glycopeptide antibiotic LY264826 with N-alkyl-linked substitutions on the epivancosamine sugar are active against glycopeptide-resistant enterococci. Six compounds representing our most active series were evaluated for activity against antibiotic-resistant, gram-positive pathogens. For Enterococcus faecium and E. faecalis resistant to both vancomycin and teicoplanin, the MICs of the six semisynthetic compounds for 90% of the strains tested were 1 to 4 micrograms/ml, compared with 2,048 micrograms/ml for vancomycin and 256 micrograms/ml for LY264826. For E. faecium and E. faecalis resistant to vancomycin but not teicoplanin, the MICs were 0.016 to 1 micrograms/ml, compared with 64 to 1,024 micrograms/ml for vancomycin. The compounds were highly active against vancomycin-susceptible enterococci and against E. gallinarum and E. casseliflavus and showed some activity against isolates of highly vancomycin-resistant leuconostocs and pediococci. The MICs for 90% of the strains of methicillin-resistant Staphylococcus aureus tested were typically 0.25 to 1 micrograms/ml, compared with 1 microgram/ml for vancomycin. Against methicillin-resistant S. epidermidis MICs ranged from 0.25 to 2 micrograms/ml, compared with 1 to 4 micrograms/ml for vancomycin and 4 to 16 micrograms/ml for teicoplanin. The spectrum of these new compounds included activity against teicoplanin-resistant, coagulase-negative staphylococci. The compounds exhibited exceptional potency against pathogenic streptococci, with MICs of < or = 0.008 microgram/ml against Streptococcus pneumoniae, including penicillin-resistant isolates. In in vivo studies with a mouse infection model, the median effective doses against a challenge by S. aureus, S. pneumoniae, or S. pyogenes were typically 4 to 20 times lower than those of vancomycin. Overall, these new glycopeptides, such as LY307599 and LY333328, show promise for use as agents against resistant enterococci, methicillin-resistant S. aureus, and penicillin-resistant pneumococci.

Reductive alkylation of glycopeptide antibiotics: synthesis and antibacterial activity.

Reductive alkylation of the A82846 family of glycopeptide antibiotics has the potential of producing seven products. N-Alkylation of the disaccharide amino function can be accomplished selectively, and offers the greatest increase in antibacterial activity. Products resulting from N-alkylation of LY264826 (A82846B) provide the most potent derivatives as compared to other members of this class of antibiotics. Two of these derivatives, LY307599 and LY333328 are approximately 500 times more active than vancomycin against vancomycin-resistant enterococci.

Activities of the semisynthetic glycopeptide LY191145 against vancomycin-resistant enterococci and other gram-positive bacteria.

LY191145 is the prototype of a series of compounds with activities against vancomycin-resistant enterococci derived by modification of the glycopeptide antibiotic LY264826. LY191145 had MICs for vancomycin- and teicoplanin-resistant enterococci of < or = 4 micrograms/ml for 50% of isolates and < or = 16 micrograms/ml for 90% of isolates. Its MICs for vancomycin-resistant, teicoplanin-susceptible enterococci were 1 to 8 micrograms/ml. LY191145 retains the potent activities of its parent compound against staphylococci and streptococci. In vivo studies in a mouse infection model confirmed these activities. This compound indicates the potential of semisynthetic glycopeptides as agents against antibiotic-resistant gram-positive bacteria.

Courageous leaders. The integral force behind organizational excellence.

For more than a decade, Total Quality Management (TQM) has been used as a powerful instrument in shaping the competitive strategies of businesses, and producing quality products and services has become the credo of firms trying to defend or expand their markets. During this time, we have come to realize that without effective leadership no quality program can succeed. That is why the quality guru, W. Edwards Deming, refused to work in any organization unless he could begin with the CEO. That is why the first criterion examined for the Malcolm Baldridge National Quality Award is leadership. Focusing on quality will not guarantee success in today's rapidly changing markets. Increasingly discriminating consumers have come to expect quality in the products and services they buy, and businesses that fail to deliver it will not survive. In a very real sense, quality performance is the price you must pay simply to play the game. But consumers want more, and satisfying their expectations will determine tomorrow's winners and losers. Building organizations capable of producing superior results that consistently meet the needs of customers is the responsibility of leaders. For this reason, leaders are more important today than they have ever been before. Leaders in successful businesses must show the way for their employees by nurturing "cultures" that encourage and reward superior performance and by exhibiting personal characteristics that inspire excellence. Great leaders possess three crucial characteristics--vision, strong values and beliefs, and the courage to do the job despite seemingly insurmountable obstacles. These characteristics make the difference between excellence and "business as usual."