Non-Essential Role for TLR2 and Its Signaling Adaptor Mal/TIRAP in Preserving Normal Lung Architecture in Mice

Myeloid differentiation factor 88 (MyD88) and MyD88-adaptor like (Mal)/Toll-interleukin 1 receptor domain containing adaptor protein (TIRAP) play a critical role in transducing signals downstream of the Toll-like receptor (TLR) family. While genetic ablation of the TLR4/MyD88 signaling axis in mice leads to pulmonary cell death and oxidative stress culminating in emphysema, the involvement of Mal, as well as TLR2 which like TLR4 also signals via MyD88 and Mal, in the pathogenesis of emphysema has not been studied. By employing an in vivo genetic approach, we reveal here that unlike the spontaneous pulmonary emphysema which developed in Tlr4^−/− mice by 6 months of age, the lungs of Tlr2^−/− mice showed no physiological or morphological signs of emphysema. A more detailed comparative analysis of the lungs from these mice confirmed that elevated oxidative protein carbonylation levels and increased numbers of alveolar cell apoptosis were only detected in Tlr4^−/− mice, along with up-regulation of NADPH oxidase 3 (Nox3) mRNA expression. With respect to Mal, the architecture of the lungs of Mal^−/− mice was normal. However, despite normal oxidative protein carbonylation levels in the lungs of emphysema-free Mal^−/− mice, these mice displayed increased levels of apoptosis comparable to those observed in emphysematous Tlr4^−/− mice. In conclusion, our data provide in vivo evidence for the non-essential role for TLR2, unlike the related TLR4, in maintaining the normal architecture of the lung. In addition, we reveal that Mal differentially facilitates the anti-apoptotic, but not oxidant suppressive, activities of TLR4 in the lung, both of which appear to be essential for TLR4 to prevent the onset of emphysema.     

Quantitative variability of 342 plasma proteins in a human twin population

The degree and the origins of quantitative variability of most human plasma proteins are largely unknown. Because the twin study design provides a natural opportunity to estimate the relative contribution of heritability and environment to different traits in human population, we applied here the highly accurate and reproducible SWATH mass spectrometry technique to quantify 1,904 peptides defining 342 unique plasma proteins in 232 plasma samples collected longitudinally from pairs of monozygotic and dizygotic twins at intervals of 2–7 years, and proportioned the observed total quantitative variability to its root causes, genes, and environmental and longitudinal factors. The data indicate that different proteins show vastly different patterns of abundance variability among humans and that genetic control and longitudinal variation affect protein levels and biological processes to different degrees. The data further strongly suggest that the plasma concentrations of clinical biomarkers need to be calibrated against genetic and temporal factors. Moreover, we identified 13 cis‐SNPs significantly influencing the level of specific plasma proteins. These results therefore have immediate implications for the effective design of blood‐based biomarker studies.     

Quirks of dye nomenclature. 6. Malachite green

Malachite green was discovered independently by two researchers in Germany in the 19^th century and found immediate employment as a dye and a pigment. Subsequently, other uses, such as staining biological specimens, emerged. A much later application was the control of fungal and protozoan infections in fish, for which the dye remains popular, although illegal in many countries owing to a variety of toxicity problems. In solution, malachite green can exist as five different species depending on the pH. The location of the positive charge of the colored cation on a carbon atom or a nitrogen atom is still debated. The original names of this dye, and their origins, are briefly surveyed.     

Inhibition of neutrophil oxidative burst and granule secretion by Wortmannin: Potential role of MAP kinase and renaturable kinases

Exposure of neutrophils to a variety of agonists including soluble chemoattractant peptides and cytokines results in degranulation and activation of the oxidative burst (effector functions) that are required for bacterial killing. At present, the signaling pathways regulating these important functions are incompletely characterized. Mitogen-activated protein (MAP) kinases (MAPK) as well as members of a family of “renaturable kinases” are rapidly activated in neutrophils in response to diverse physiological agonists, suggesting that they may regulate cell activation. Antagonists of phosphatidyl inositol-3-(OH) kinase (PI3-kinase) such as wortmannin (Wtmn) inhibit these effector responses as well as certain of the above-mentioned kinases, leading to the suggestion that these enzymes lie downstream of PI3-kinase in the pathway regulating the oxidative burst and granule secretion. However, an apparent discrepancy exists in that, while virtually obliterating activity of PI3-kinase and the oxidase at low concentrations (ID₅₀ < 20 nM), Wtmn has only variable inhibitory effects on MAPK even at substantially higher concentrations (75–100 nM). This raises the possibility that the inhibitory effects of Wtmn are mediated via other enzyme systems. The purpose of the current study was therefore to compare the effects of Wtmn on PI3-kinase activity and on the chemoattractant-activated kinases, and to determine the potential relationship of these pathways to microbicidal responses. In human neutrophils, both the oxidative burst and granule secretion induced by fMLP were inhibited by Wtmn but at markedly different concentrations: the oxidative burst was inhibited with an ID₅₀ of <5 nM while granule secretion was only partially inhibited at concentrations exceeding 75 nM. Activation of both MEK-1 and MAPK in response to fMLP was only partially inhibited by high doses of Wtmn (ID₅₀ of > 100 nM and ≡75 nM, respectively). In contrast, Wtmn potently inhibited fMLP-induced activation of the 63 and 69 kDa renaturable kinases (ID₅₀ ≡ 5–10 nM). We speculate that the renaturable kinases may be involved in the regulation of the oxidative burst, whereas the MAPK pathway may play a role in other neutrophil functions such as granule secretion. J. Cell. Physiol. 172:94–108, 1997. © 1997 Wiley-Liss, Inc.     

Structural and functional analysis of the GABARAP interaction motif (GIM)

Through the canonical LC3 interaction motif (LIR), [W/F/Y]‐X₁‐X₂‐[I/L/V], protein complexes are recruited to autophagosomes to perform their functions as either autophagy adaptors or receptors. How these adaptors/receptors selectively interact with either LC3 or GABARAP families remains unclear. Herein, we determine the range of selectivity of 30 known core LIR motifs towards individual LC3s and GABARAPs. From these, we define a I nteraction 相似文献   

EFFECT OF VITAMIN E SUPPLEMENTATION AND PARTIAL SUBSTITUTION OF POLY- WITH MONO-UNSATURATED FATTY ACIDS IN PIG DIETS ON MUSCLE,AND MICROSOME EXTRACT a-TOCOPHEROL CONCENTRATION AND LIPID OXIDATION

The experiment was organized in a 3×2 factorial arrangement with three dietary fat blends and a basal (20 mg kg^?1 diet) or supplemented (220 mg kg^?1) level of α-tocopheryl acetate. Dietary vitamin E and monounsaturated to polyunsaturated fatty acid ratio (dietary MUFA/PUFA) affected muscle α-tocopherol concentration (α-tocopherol [log μg g^?1]=0.18 (±0.105)+0.0034 (±0.0003)·dietary α-tocopherol [mg kg^?1 diet] (P<0.0001)+0.39 (±0.122)·dietary MUFA/PUFA (P<0.0036)). An interaction between dietary α-tocopherol and dietary MUFA/PUFA exists for microsome α-tocopherol concentration (α-tocopherol [log μg g^?1]=1.14 (±0.169) (P<0.0001)+0.0056 (±0.00099)·dietary α-tocopherol [mg kg^?1 diet] (P<0.0001)+0.54 (±0.206)·dietary MUFA/PUFA (P<0.0131)?0.0033 (±0.0011)·dietary α-tocopherol [mg kg^?1)]×dietary MUFA/PUFA (P<0.0067)), and hexanal concentration in meat (hexanal [ng·g^?1]=14807.9 (±1489.8)?28.8 (±10.6) dietary α-tocopherol [mg·kg^?1] (P<0.01)?8436.6 (±1701.6)·dietary MUFA/PUFA (P<0.001)+24.0 (±11.22)·dietary α-tocopherol·dietary MUFA/PUFA (P<0.0416)). It is concluded that partial substitution of dietary PUFA with MUFA lead to an increase in the concentration of α-tocopherol in muscle and microsome extracts. An interaction between dietary α-tocopherol and fatty acids exists, in which at low level of dietary vitamin E inclusion, a low MUFA/PUFA ratio leads to a reduction in the concentration of α-tocopherol in microsome extracts and a concentration of hexanal in meat above the expected values.     

Quirks of dye nomenclature. 5. Rhodamines

Rhodamines were first produced in the late 19^th century, when they constituted a new class of synthetic dyes. These compounds since have been used to color many things including cosmetics, inks, textiles, and in some countries, food products. Certain rhodamine dyes also have been used to stain biological specimens and currently are widely used as fluorescent probes for mitochondria in living cells. The early history and current biological applications are sketched briefly and an account of the ambiguities, complications and confusions concerning dye identification and nomenclature are discussed.     

Application of an optimized marker amplification protocol in immunohistochemistry — a valuable tool for visualizing antigenic sites of low signal strength or sparse distribution

Amplification of immunohistochemical markers received considerable attention during the 1980s and 1990s. The amplification approach was largely abandoned following the development of antigen retrieval and reporter amplification techniques, because the latter were incorporated more easily into high throughput automated procedures in industrial and diagnostic laboratories. There remain, however, a number of instances where marker amplification still has much to offer. Consequently, we examined experimentally the utility of an optimized marker amplification technique in diagnostically relevant tissue where either the original signal strength was low or positive sites were visible, but sparsely distributed. Marker amplification in the former case not only improved the visibility of existing positive sites, but also revealed additional sites that previously were undetectable. In the latter case, positive sites were rendered more intense and therefore more easily seen during low magnification examination of large areas of tissue.     

Antigen-induced airway inflammation in the Brown Norway rat results in airway smooth muscle hyperplasia.

Asthma is characterized by chronic airways inflammation, airway wall remodeling, and airway hyperresponsiveness (AHR). An increase in airway smooth muscle has been proposed to explain a major part of AHR in asthma. We have used unbiased stereological methods to determine whether airway smooth muscle hyperplasia and AHR occurred in sensitized, antigen-challenged Brown Norway (BN) rats. Ovalbumin (OA)-sensitized BN rats chronically exposed to OA aerosol displayed airway inflammation and a modest level of AHR to intravenously administered ACh 24 h after the last antigen challenge. However, these animals did not show an increase in smooth muscle cell (SMC) number in the left main bronchus, suggesting that short-lived inflammatory mechanisms caused the acute AHR. In contrast, 7 days after the last aerosol challenge, there was a modest increase in SMC number, but no AHR to ACh. Addition of FCS to the chronic OA challenge protocol had no effect on the degree of inflammation but resulted in a marked increase in both SMC number and a persistent (7-day) AHR. These results raise the possibility that increases in airway SMC number rather than, or in addition to, chronic inflammation contribute to the persistent AHR detected in this model.