5'-Oligonucleotides as the acid-soluble products of the ATP-dependent DNase form Escherichia coli

The acid-soluble products of the ATP-dependent DNase action on DNA were identified to be a mixture of 5′-oligonucleotides the average chain length of which is approximately 3, ranging from mono- to pentanucleotides.     

Enalapril and treadmill running reduce adiposity,but only the latter causes adipose tissue browning in mice

This study investigated whether regulation of the renin–angiotensin system (RAS) by enalapril and/or aerobic exercise training (AET) causes browning of the subcutaneous white adipose tissue (sWAT). C57BL/6 mice were fed either a standard chow or a high‐fat (HF) diet for 16 weeks. At Week 8, HF‐fed animals were divided into sedentary (HF), enalapril (HF‐E), AET (HF‐T), and enalapril plus AET (HF‐ET) groups. Subsequently, sWAT was extracted for morphometry, determination of RAS expression, and biomarkers of WAT browning. The HF group displayed adipocyte hypertrophy and induction of the classical RAS axis. Conversely, all interventions reduced adiposity and induced the counterregulatory RAS axis. However, only AET raised plasma irisin, increased peroxisome proliferator‐activated receptor‐γ coactivator‐1α, and uncoupling protein‐1 levels, and the expression of PR‐domain containing 16 in sWAT. Therefore, we concluded that AET‐induced sWAT browning was independent of the counterregulatory axis shifting of RAS in HF diet‐induced obesity.     

Genetic control of the B cell response to LPS: opposing effects in peritoneal versus splenic B cell populations

Lipopolysaccharide (LPS) from gram-negative bacteria activates B cells, enabling them to proliferate and differentiate into plasma cells. This response is critically dependent on the expression of TLR4; but other genes, such as RP105 and MHC class II, have also been shown to contribute to B cell LPS response. Here, we have evaluated the role of genetic control of the B cell response to LPS at the single cell level. We compared the response to LPS of peritoneal cavity (PEC) and splenic B cells on the BALB/c genetic background (LPS-low responder) to those on the C57BL/6J background (LPS-high responder) and their F1 progeny (CB6F1). Both PEC and splenic B cells from B6 exhibited 100% clonal growth in the presence of LPS; whereas, BALB/c PEC and splenic B cells achieved only 50% and 23% clonal growth, respectively. Adding CpG to the LPS stimulus pushed PEC B cell clonal growth in the low responder strain BALB/c up to 90%, showing that the nonresponse to LPS is a specific effect. Surprisingly, PEC B cells on the F1 background behaved as high responders, while splenic B cells behaved as low responders to LPS. The data presented here reveals a previous unsuspected behavior in the genetic control of the B cell response to LPS with an opposing impact in splenic versus peritoneal cavity B cells. These results suggest the existence of an, as yet, unidentified genetic factor exclusively expressed by coelomic B cells that contributes to the control of the LPS signaling pathway in the B lymphocyte.     

Recent advances in immobilization strategies for glycosidases

Glycans play important biological roles in cell‐to‐cell interactions, protection against pathogens, as well as in proper protein folding and stability, and are thus interesting targets for scientists. Although their mechanisms of action have been widely investigated and hypothesized, their biological functions are not well understood due to the lack of deglycosylation methods for large‐scale isolation of these compounds. Isolation of glycans in their native state is crucial for the investigation of their biological functions. However, current enzymatic and chemical deglycosylation techniques require harsh pretreatment and reaction conditions (high temperature and use of detergents) that hinder the isolation of native glycan structures. Indeed, the recent isolation of new endoglycosidases that are able to cleave a wider variety of linkages and efficiently hydrolyze native proteins has opened up the opportunity to elucidate the biological roles of a higher variety of glycans in their native state. As an example, our research group recently isolated a novel Endo‐β‐N‐acetylglucosaminidase from Bifidobacterium longum subsp. infantis ATCC 15697 (EndoBI‐1) that cleaves N‐N′‐diacetyl chitobiose moieties found in the N‐linked glycan (N‐glycan) core of high mannose, hybrid, and complex N‐glycans. This enzyme is also active on native proteins, which enables native glycan isolation, a key advantage when evaluating their biological activities. Efficient, stable, and economically viable enzymatic release of N‐glycans requires the selection of appropriate immobilization strategies. In this review, we discuss the state‐of‐the‐art of various immobilization techniques (physical adsorption, covalent binding, aggregation, and entrapment) for glycosidases, as well as their potential substrates and matrices. © 2016 American Institute of Chemical Engineers Biotechnol. Prog., 33:104–112, 2017     

Characterization of the intracellular Ca(2+) pools involved in the calcium homeostasis in Herpetomonas sp. promastigotes

Trypanosomatids of the genus Herpetomonas comprises monoxenic parasites of insects that present pro- and opisthomastigotes forms in their life cycles. In this study, we investigated the Ca(2+) transport and the mitochondrial bioenergetic of digitonin-permeabilized Herpetomonas sp. promastigotes. The response of promastigotes mitochondrial membrane potential to ADP, oligomycin, Ca(2+), and antimycin A indicates that these mitochondria behave similarly to vertebrate and Trypanosoma cruzi mitochondria regarding the properties of their electrochemical proton gradient. Ca(2+) transport by permeabilized cells appears to be performed mainly by the mitochondria. Unlike T. cruzi, it was not possible to observe Ca(2+) release from Herpetomonas sp. mitochondria, probably due to the simultaneous Ca(2+) uptake by the endoplasmic reticulum. In addition, a vanadate-sensitive Ca(2+) transport system, attributed to the endoplasmic reticulum, was also detected. Nigericin (1 microM), FCCP (1 microM), or bafilomycin A(1) (5 microM) had no effect on the vanadate-sensitive Ca(2+) transport. These data suggest the absence of a Ca(2+) transport mediated by a Ca(2+)/H(+) antiport. No evidence of a third Ca(2+) compartment with the characteristics of the acidocalcisomes described by A. E. Vercesi et al. (1994, Biochem. J. 304, 227-233) was observed. Thapsigargin and IP(3) were not able to affect the vanadate-sensitive Ca(2+) transport. Ruthenium red was able to inhibit the Ca(2+) uniport of mitochondria, inducing a slow mitochondrial Ca(2+) efflux, compatible with the presence of a Ca(2+)/H(+) antiport. Moreover, this efflux was not stimulated by the addition of NaCl, which suggests the absence of a Ca(2+)/Na(+) antiport in mitochondria.     

Expression of functional TLR4 confers proinflammatory responsiveness to Trypanosoma cruzi glycoinositolphospholipids and higher resistance to infection with T. cruzi

TLRs function as pattern recognition receptors in mammals and play an essential role in the recognition of microbial components. We found that the injection of glycoinositolphospholipids (GIPLs) from Trypanosoma cruzi into the peritoneal cavity of mice induced neutrophil recruitment in a TLR4-dependent manner: the injection of GIPL in the TLR4-deficient strain of mice (C57BL/10ScCr) caused no inflammatory response. In contrast, in TLR2 knockout mice, neutrophil chemoattraction did not differ significantly from that seen in wild-type controls. GIPL-induced neutrophil attraction and MIP-2 production were also severely affected in TLR4-mutant C3H/HeJ mice. The role of TLR4 was confirmed in vitro by testing genetically engineered mutants derived from TLR2-deficient Chinese hamster ovary (CHO)-K1 fibroblasts that were transfected with CD14 (CHO/CD14). Wild-type CHO/CD14 cells express the hamster TLR4 molecule and the mutant line, in addition, expresses a nonfunctional form of MD-2. In comparison to wild-type cells, mutant CHO/CD14 cells failed to respond to GIPLs, indicating a necessity for a functional TLR4/MD-2 complex in GIPL-induced NF-kappaB activation. Finally, we found that TLR4-mutant mice were hypersusceptible to T. cruzi infection, as evidenced by a higher parasitemia and earlier mortality. These results demonstrate that natural resistance to T. cruzi is TLR4 dependent, most likely due to TLR4 recognition of their GIPLs.     

Mechanisms for increasing stroke volume during static exercise with fixed heart rate in humans

Nóbrega, Antonio C. L., Jon W. Williamson, Jorge A. Garcia, and Jere H. Mitchell. Mechanisms for increasing stroke volume during static exercise with fixed heart rate in humans. J. Appl. Physiol. 83(3): 712-717, 1997.Ten patients with preserved inotropic function having adual-chamber (right atrium and right ventricle) pacemaker placed forcomplete heart block were studied. They performed static one-leggedknee extension at 20% of their maximal voluntary contraction for 5 minduring three conditions: 1)atrioventricular sensing and pacing mode [normal increase in heart rate (HR; DDD)], 2) HRfixed at the resting value (DOO-Rest; 73 ± 3 beats/min), and3) HR fixed at peak exercise rate(DOO-Ex; 107 ± 4 beats/min). During control exercise (DDD mode),mean arterial pressure (MAP) increased by 25 mmHg with no change instroke volume (SV) or systemic vascular resistance. During DOO-Rest andDOO-Ex, MAP increased (+25 and +29 mmHg, respectively) because of aSV-dependent increase in cardiac output (+1.3 and +1.8 l/min,respectively). The increase in SV during DOO-Rest utilized acombination of increased contractility and the Frank-Starling mechanism(end-diastolic volume 118-136 ml). However, during DOO-Ex, agreater left ventricular contractility (end-systolic volume 55-38ml) mediated the increase in SV.