Adsorption from black tea and red wine onto in vitro salivary pellicles studied by ellipsometry

The adsorption of black tea and red wine components onto a pellicle-like protein layer formed in vitro by adsorption from whole unstimulated saliva on hydroxyapatite discs were studied by in situ ellipsometry. It was found that components from black tea readily adsorbed to the pellicle. Subsequent exposure to saliva led to further adsorption of salivary components to give an overall increase in the amounts adsorbed. The amounts adsorbed increased still further following a third tea and saliva exposure. Components of red wine gave significantly greater amounts of adsorption to the pellicle than black tea. The adsorption of components of black tea gave a concomitant increase in colour or stain as measured by a reflectance chromameter. In all cases, the black tea- and red wine-modified pellicles were not eluted by either phosphate buffer or sodium dodecyl sulphate (SDS) rinses. Thus, black tea and red wine components have been shown to have a profound effect on in vitro pellicle maturation, causing thickened layers of stained material to build up, which are not readily removed.     

可溶性凝集素样氧化型低密度脂蛋白受体 -1在急性缺血性脑卒中表达及与静脉溶栓预后和静脉溶栓后脑出血转化的关系

目的探讨可溶性凝集素样氧化型低密度脂蛋白受体 -1(sLOX-1)在急性缺血性脑卒中( AIS)表达及与静脉溶栓预后和静脉溶栓后脑出血转化的关系。方法选取 2017年 3月至 2020年 6月南京市高淳人民医院 AIS病人 104例,酶联免疫吸附法(ELISA)法测定血清 sLOX-1水平,比较 sLOX-1表达高低与病人临床资料的关系,分析 sLOX-1与 AIS病人静脉溶栓预后和静脉溶栓后脑出血转化的关系。结果相比于低 sLOX-1表达组,高 sLOX-1表达组病症更加严重,美国国立卫生院卒中量表(NHISS)评分显著增大,白细胞显著升高,肾小球滤过率估算值( eGFR)尿酸(UA)显著降低(均 P<0.05);行重组组织型纤溶酶原( rt-PA)静脉溶栓治疗预后不良组 sLOX-1表达显著高于预后良好组［(2、.18±0.71)比( 1.67±0.53)］;多因素 logistic回归模型结果发现经过年龄、溶栓时间、 NHISS评分、 eGFR等校正过后, sLOX-1［OR=3.83,95%CI:(1.77,5.12)P=0.007］仍然是行 rt-PA静脉溶栓 AIS病人预后的独立危险因素,经过年龄、 NHISS评分、溶栓时间等校正过后, sLOX-1［OR=1.84,9,5%CI:(1.12,2.54)P= 0.020］也仍然是 AIS病人发生脑出血性转化( HT)的独立危险因素。结论血清 sLOX-1是行 rt-PA静脉溶栓 AIS病人预后立危险因素,也是 AIS病人发生脑 HT的独立危险因素。     

The complement system in primary Sjögren’s syndrome: the expression of certain cascade and regulatory proteins in labial salivary glands – observational study

IntroductionThe complement cascade and regulatory proteins are involved in the pathogenesis of the Sjögren’s syndrome and other autoimmune diseases. The complement activation via the alternative pathway was recognized as a major pathogenic mechanism in autoimmune conditions. The aim of this study was to assess expression of complement cascade components and regulatory proteins in minor salivary glands in patients with primary Sjögren’s syndrome (pSS).Materials and methodsThe expression of C1q and C5b-9 – membrane attack complex and regulatory proteins such as: membrane cofactor protein (MCP), decay-accelerating factor (DAF) and protectin were examined using immunochemistry method in specimens from biopsy of minor salivary glands in pSS patients. The biopsy material was obtained from 20 pSS patients, 5 patients with non-specific sialadenitis and from 5 patients with suspicion of dryness syndrome without sialadenitis confirmation.ResultsNone of the examined samples showed the expression of C1q or the effector C5b-9. Membrane cofactor protein expression was lower in pSS group than in both non-specific sialadenitis and noninflamed salivary glands. The inflammatory cells in pSS samples partially expressed MCP. There were differences in the sites and intensity of membrane protectin expression exclusively on the luminal surfaces in pSS; on the luminal and, partially, antiluminal surface in non-specific inflammation, and on the entire cell surface in unaffected salivary glands. There were no DAF expression in salivary gland tissue in biopsy specimens in all studied subjects.ConclusionsThe study demonstrated the absence of complement-cascade proteins (C1q, MAC) in the salivary glands of pSS patients, which may indicated a lack of local complement activation via the classical pathway and the observed gland tissue damage being due to a mechanism other than MAC-induced cytolysis. The differences in the expression of complement regulatory proteins between pSS, non-specific sialadenitis, and normal salivary glands may indicate that alternative functions of these regulatory proteins may be of greater significance in pSS. Low MCP expression in pSS in comparison with non-specific sialadenitis and normal salivary glands, may suggest altered modulation of cell-mediated immunity in pSS. The differences in the location and intensity of protectin (CD59) expression indicates a possibility of reducing the proinflammatory effect of protectin in pSS.     

Tailored use of belatacept in adolescent kidney transplantation

Adolescent transplant recipients are at risk for nonadherence, development of de novo donor‐specific antibody (dnDSA), and allograft loss. Belatacept, a selective T cell costimulatory blocker, is associated with reduced dnDSA, improved renal function, and prolonged allograft survival when compared to calcineurin inhibitor‐based regimens in adults; however, its use in children is scant. Three adolescents were initiated on belatacept between August 2017 and September 2018 at the time of kidney transplantation. Selection criteria included age ≥ 14 and EBV IgG + serostatus. Intraoperative alemtuzumab and methylprednisolone were given as induction therapy. Tailored maintenance therapy included steroid‐free belatacept and sirolimus for two patients. One patient was initially maintained steroid‐free on belatacept and belimumab, an inhibitor of B cell activating factor to treat concurrent systemic lupus erythematous; steroids were added subsequently. Renal function, biopsy‐proven rejection, dnDSA, allograft survival, infection, nonadherence, and proteinuria were monitored. Renal function was 86, 73, 52 mL/min/1.73 m² at 20, 20, and 8 months, respectively. There was 100% adherence to therapy and no development of dnDSA. All patients had treatable infections. One developed steroid‐responsive acute cellular rejection. Belatacept‐based regimens can be tailored for adolescent recipients with good short‐term clinical outcomes.     

Zika virus: what we need to know?

Zika virus is one of the emerging viruses and is of significant threat to human health globally. It is a mosquito borne flavivirus similar to dengue, yellow fever, and West Nile viruses. It was reported about 5 decades ago and then it spreads to different parts of the world. Large outbreaks were reported on Yap Islands in 2007. Now it has gained wide attention globally by health communities. Major vector for virus transmission is Aedes aegypti mosquito. ZIKV infection is mostly asymptomatic but it is also responsible to cause mild influenza like illness to serious manifestations. There is no specific anti‐viral treatment is available for ZIKV infection. The virus disseminates very fast due to which it possesses a serious threat especially in those areas where there is lack of specific immunity against virus. Little knowledge is available on its transmission and pathogenicity. Although virus was discovered years ago but its genomic structure is not clearly understood yet. In this review we focus on the current knowledge of epidemiology of ZIKV, its transmission, its structural biology, different aspects of diagnosis and diagnostic challenges as well as highlighted appropriates antiviral drugs and vaccines regarding treatment.     

Autophagy limits activation of the inflammasomes

Inflammasomes are multiprotein complexes that control the maturation and production of interleukin-1 family members and play crucial roles in host defense against pathogens. However, dysregulated activation of inflammasomes is associated with intense inflammation, leading to the development of inflammatory diseases. Therefore, inflammasomes must be activated at a proper strength to protect against infection and avoid tissue damage. Recent studies have highlighted the cross-talk between inflammasome activation and autophagy, the cellular machinery associated with the degradation of intracellular components and maintenance of cellular homeostasis. Notably, deficiencies in autophagy-related proteins induce the aberrant activation of inflammasomes, causing severe tissue damage. In contrast, autophagy inducers ameliorate symptoms of inflammasome-related diseases. In this review, we discuss recent advances in the involvement of autophagy in regulating inflammasomes activation and in the development of inflammatory diseases.     

Global structural changes of an ion channel during its gating are followed by ion mobility mass spectrometry

Mechanosensitive ion channels are sensors probing membrane tension in all species; despite their importance and vital role in many cell functions, their gating mechanism remains to be elucidated. Here, we determined the conditions for releasing intact mechanosensitive channel of large conductance (MscL) proteins from their detergents in the gas phase using native ion mobility–mass spectrometry (IM-MS). By using IM-MS, we could detect the native mass of MscL from Escherichia coli, determine various global structural changes during its gating by measuring the rotationally averaged collision cross-sections, and show that it can function in the absence of a lipid bilayer. We could detect global conformational changes during MscL gating as small as 3%. Our findings will allow studying native structure of many other membrane proteins.One of the best candidates to explore the gating of mechanosensitive channels is the mechanosensitive channel of large conductance (MscL) from Escherichia coli. The crystal structure of MscL in its closed/nearly closed state from Mycobacterium tuberculosis revealed this channel as a homopentamer (1). Each subunit has a cytoplasmic N- and C-terminal domain as well as two α-helical transmembrane (TM) domains, TM1 and TM2, which are connected by a periplasmic loop. The five TM1 helices form the pore and the more peripheral TM2 helices interact with the lipid bilayer.MscL detects changes in membrane tension invoked by a hypoosmotic shock and couples the tension sensing directly to large conformational changes (1, 2). On the basis of a large body of structural and theoretical data, numerous gating models of MscL have been proposed (3–9). These models agree upon (i) the hydrophobic pore constriction of the channel and (ii) the channel opens by an iris-like rotation—i.e., a tilting and outward movement of transmembrane helices that make the channel wider and shorter (5). This mechanism is supported by patch-clamp (10), disulfide cross-linking (11), FRET spectroscopy (12), and site-directed spin labeling EPR experiments (6, 7), as well as computational studies (13–15). So far, direct experimental results have only been observed for short-range local structural changes, and no measure of the overall global structural changes during channel gating have been reported. Because there is no crystal structure available for the open MscL channel, elucidating overall global structural changes from the onset of channel activation is of utmost importance for our understanding of the gating mechanism of mechanosensitive channels. Here, we provide direct experimental evidence for the key areal changes occurring during channel gating by combining our ability to activate MscL in a controlled manner to different subopen states (16) with a native ion mobility-mass spectrometry (IM-MS) approach.     

Accounting for a mirror-image conformation as a subtle effect in protein folding

By using local (free-energy profiles along the amino acid sequence and ¹³C^α chemical shifts) and global (principal component) analyses to examine the molecular dynamics of protein-folding trajectories, generated with the coarse-grained united-residue force field, for the B domain of staphylococcal protein A, we are able to (i) provide the main reason for formation of the mirror-image conformation of this protein, namely, a slow formation of the second loop and part of the third helix (Asp29–Asn35), caused by the presence of multiple local conformational states in this portion of the protein; (ii) show that formation of the mirror-image topology is a subtle effect resulting from local interactions; (iii) provide a mechanism for how protein A overcomes the barrier between the metastable mirror-image state and the native state; and (iv) offer a plausible reason to explain why protein A does not remain in the metastable mirror-image state even though the mirror-image and native conformations are at least energetically compatible.To perform their functions in living organisms, most proteins must fold from unfolded polypeptides into their functional, unique 3D structures. Understanding protein-folding mechanisms is crucial because misfolded proteins can cause many diseases, including neurodegenerative diseases (1) such as Alzheimer’s, Parkinson, and Huntington diseases. From theoretical and conceptual points of view, it has been suggested that a native protein exists in a thermodynamically stable state with its surroundings (2) and that a study of free-energy landscapes (FELs) holds the key to understanding how proteins fold and function (3, 4).The native structures of some proteins contain a high degree of symmetry that, in addition to the native structure, allows the existence of another, energetically very close to the native conformation, a native-like “mirror-image” structure. One of the representatives of such symmetrical proteins is the 10- to 55-residue fragment of the B domain of staphylococcal protein A [Protein Data Bank (PDB) ID: 1BDD, a three-α-helix bundle] (5). Protein A has been the subject of extensive theoretical (6–18) and experimental (19–23) studies because of its small size, fast folding kinetics, and biological importance. However, the mirror-image topology has never been a subject for discussion except for the earlier work by Olszewski et al. (7) and recent work by Noel et al. (24). The reason for this might be that it has never been detected experimentally and it was observed only in some theoretical studies (7–9, 12, 13, 15, 17, 18, 24) with different force fields. It is of interest to determine how realistic the mirror-image conformation is. Is it an artifact of the simulations or is it a conformation difficult to observe experimentally? Noel et al. (24) showed that the native and mirror-image structures have a similar enthalpic stability and are thermodynamically competitive and that the mirror image can be considered not just a computational annoyance, but as a real conformation competing with the native structure. Moreover, the mirror-image conformation is more entropically favorable than the native conformation (24). By making multiple mutations in the hydrophobic core and the first loop region, Olszewski et al. (7) found that the change in the handedness of the first loop induced by the mutations, the burial of the N cap of the second helix, and repacking of the hydrophobic core are responsible for formation of the mirror-image conformation. However, at the end, the authors stated: “… Whether the conclusion about the possible importance of turns in defining the global topology holds in general or is just specific to the three-helix bundles analyzed here requires additional investigation....” (ref. 7, p. 298).The difficulties for experiments to detect the mirror-image topology arise because the secondary structures of the mirror-image and the native conformation are identical and the native-contact interactions are similar in both conformations (details in Fig. S1 and SI Native and Mirror-Image Structures of Protein A). Hence, with an experimental technique such as circular dichroism, used to estimate the fraction of secondary-structure content, it is almost impossible to distinguish the mirror-image structure from the native structure. It would have been desirable if the mirror-image conformation and its evolution to the native structure could be detected by NMR spectroscopy. Nevertheless, by using local [¹³C^α chemical shift (25) and free-energy profiles (FEPs) along the amino acid sequence (26–28)] and global [principal component (PC) (29)] analyses (SI Materials and Methods), we examined molecular dynamics (MD) trajectories of protein A, generated with the coarse-grained united-residue (UNRES) force field (27, 30–32) (Fig. S2 and SI Materials and Methods). These analyses of the MD trajectories, in which folding from a fully unfolded conformation occurs either almost instantly or through a metastable state formed by the mirror-image topology, enabled us to elucidate the origin of the formation of a mirror-image topology and how the protein emerges from the kinetic trap and folds to the native state.The results presented in this work are based on the analysis of four pairs of MD trajectories at 270 K (in each pair, one trajectory folds directly to the native state and the other folds through the metastable mirror-image state) selected from 96 MD simulations, which we carried out in a broad range of temperatures (details in Materials and Methods). The mirror-image conformation is energetically competitive with the native conformation in the studied trajectories (an illustrative example of two trajectories is in Fig. S3), and these results are in agreement with those of earlier studies (12, 24).     

肺炎链球菌细胞壁表面粘附相关蛋白的初步鉴定

通过Ｗｅｓｔｅｒｎｂｌｏｔ将肺炎链球菌细胞壁表面蛋白质转移到ＰＶＤＦ膜上，然后与生物素标记的巨噬细胞在膜上进行粘附实验，初步鉴定出六条细胞壁表面粘附相关蛋白，它们的分子量分别是２０，３０，３７，５９，６６，８５ｋＤ其生物学特性，功能有待进一步研究确定，这为进一步研究肺炎链球菌致病机理，发展新一代多价疫苗和开发抗菌药物奠定了一定的基础。     

Clinical Outcomes Following Regenerative Therapy of Non‐Contained Intrabony Defects Using a Deproteinized Bovine Bone Mineral Combined With Either Enamel Matrix Derivative or Collagen Membrane

Background: The purpose of this study is to compare clinical outcomes in the treatment of deep non‐contained intrabony defects (i.e., with ≥70% 1‐wall component and a residual 2‐ to 3‐wall component in the most apical part) using deproteinized bovine bone mineral (DBBM) combined with either enamel matrix protein derivative (EMD) or collagen membrane (CM). Methods: Forty patients with multiple intrabony defects were enrolled. Only one non‐contained defect per patient with an intrabony depth ≥3 mm located in the interproximal area of single‐ and multirooted teeth was randomly assigned to the treatment with either EMD + DBBM (test: n = 20) or CM + DBBM (control: n = 20). At baseline and after 12 months, clinical parameters including probing depth (PD) and clinical attachment level (CAL) were recorded. The primary outcome variable was the change in CAL between baseline and 12 months. Results: At baseline, the intrabony component of the defects amounted to 6.1 ± 1.9 mm for EMD + DBBM and 6.0 ± 1.9 mm for CM + DBBM sites (P = 0.81). The mean CAL gain at sites treated with EMD + DBBM was not statistically significantly different (P = 0.82) compared with CM + DBBM (3.8 ± 1.5 versus 3.7 ± 1.2 mm). No statistically significant difference (P = 0.62) was observed comparing the frequency of CAL gain ≥4 mm between EMD + DBBM (60%) and CM + DBBM (50%) or comparing the frequency of residual PD ≥6 mm between EMD + DBBM (5%) and CM + DBBM (15%) (P = 0.21). Conclusion: Within the limitations of the present study, regenerative therapy using either EMD + DBBM or CM + DBBM yielded comparable clinical outcomes in deep non‐contained intrabony defects after 12 months.