高压氧治疗对阿尔茨海默病大鼠海马内神经元凋亡及p38丝裂原蛋白激酶表达的影响

目的 评价高压氧治疗对阿尔茨海默病(alzheimer's disease,AD)大鼠海马内神经元凋亡及p38丝裂原蛋白激酶(p38 mitogenactivated protein kinases,p38MAPK)表达的影响. 方法 18只雄性SD大鼠,体重260 g～290 g,采用随机数字表法分为3组(每组6只):生理盐水组(NS组),海马区注射生理盐水;阿尔茨海默病组(AD组),海马区注射Aβ1-40;高压氧组(HBO组),海马区注射Aβ1-40,并于术后1 d～7d行每天1次的高压氧治疗.应用脑立体定位仪于大鼠海马区注射Aβ1-40制备AD模型.于术前1d和术后1、7、14、21 d行水迷宫行为学测试.于术后21 d水迷宫行为学测试结束后处死大鼠,采用苏木精-伊红染色(hematoxylin-eosin,HE)方法染色海马组织病理结构,TUNEL法标记检测凋亡细胞,免疫印迹法(Western bolt)法测定海马内p38MAPK蛋白磷酸化水平. 结果 术后21 d时,与NS组比较,AD组大鼠上台前路程和逃避潜伏期增加[(379±41)、(1 978±120) cm,(16.0±2.8)、(78.4±10.8)s](P＜0.05);与AD组比较,HBO组大鼠上台前路程和逃避潜伏期减少[(1978±120)、(1 246±96) cm,(78.4±10.8)、(51.7±6.2)s](P＜0.05).与NS组比较,AD组和HBO组大鼠海马组织细胞凋亡率[(6.3±13)％,(45.2±5.1)％]及p38MAPK蛋白磷酸化表达升高(0.16±0.06),(0.54±0.10)(P＜0.05);与AD组比较,HBO组海马组织细胞凋亡率[(45.2±5.1)％,(22.5±3.7)％]及p38 MAPK蛋白磷酸化表达下降[(0.54±0.10),(0.31±0.08)](P＜0.05).结论 高压氧治疗可有效缓解AD大鼠认知功能障碍,且能够抑制海马内神经元凋亡,其机制与降低p38MAPK的磷酸化水平有关.     

The effect of different treatment strategies on left atrial size in patients with lone paroxysmal atrial fibrillation—a prospective cohort study

INTRODUCTION: The effect of different treatment strategies of atrial fibrillation on left atrial (LA) size has not been compared in lone paroxysmal atrial fibrillation. The objective of the present study was to evaluate the evolution of LA size over time in patients who underwent different treatment interventions. METHODS AND RESULTS: Two hundred forty patients with lone paroxysmal atrial fibrillation were assigned to four groups. The circumferential pulmonary vein ablation (CPVA) group (n = 60) was treated with CPVA, segmental pulmonary vein isolation (SPVI) group (n = 60) with SPVI, AMIO group (n = 60) with amiodarone alone, and AMIO + LO group (n = 60) with amiodarone plus losartan. LA diameter was measured with transthoracic echocardiogram at baseline, 3, 6, 9, and 12 months after the interventions. In the CPVA group, LA size at third, sixth, ninth, and 12th month had a significant decrease than that at baseline and in the other three groups. LA size in patients with atrial fibrillation recurrence in the four groups was significant higher than that in patients with no atrial fibrillation recurrence (P = 0.002-0.001). CONCLUSION: The results suggested that a shortened LA size is not consistent with improved sinus rhythm maintenance. Although maintenance of sinus rhythm is not the only factor in determining shrinking or enlargement of the left atrium, inhibiting or eliminating activity of the pulmonary vein is very important for paroxysmal atrial fibrillation. Atrial fibrillation recurrence is a main factor contributing to enlargement of the LA.     

Early Childhood Gut Microbiomes Show Strong Geographic Differences Among Subjects at High Risk for Type 1 Diabetes

OBJECTIVE

Gut microbiome dysbiosis is associated with numerous diseases, including type 1 diabetes. This pilot study determines how geographical location affects the microbiome of infants at high risk for type 1 diabetes in a population of homogenous HLA class II genotypes.

RESEARCH DESIGN AND METHODS

High-throughput 16S rRNA sequencing was performed on stool samples collected from 90 high-risk, nonautoimmune infants participating in The Environmental Determinants of Diabetes in the Young (TEDDY) study in the U.S., Germany, Sweden, and Finland.

RESULTS

Study site–specific patterns of gut colonization share characteristics across continents. Finland and Colorado have a significantly lower bacterial diversity, while Sweden and Washington state are dominated by Bifidobacterium in early life. Bacterial community diversity over time is significantly different by geographical location.

CONCLUSIONS

The microbiome of high-risk infants is associated with geographical location. Future studies aiming to identify the microbiome disease phenotype need to carefully consider the geographical origin of subjects.     

胰腺癌的条件生存率

目的分析胰腺癌切除术后,在存活一定时间的基础上,评估其额外生存若干年/月的可能性。方法对2007年1月-2012年12月由该院胰腺外科专家完成的胰腺癌切除手术的所有资料进行总结分析。先通过对临床病理学变量资料的评估来预测患者的生存时间。条件生存率(CS)的计算公式是CS=S(X+Y)/Sx,x是已经存活时间,Y是额外存活时间;实际生存时间是在已经存活时间的基础上,通过个体化的方式直接计算出的。结果对于任一总体生存时间而言,CS随着已经存活时间的延长而增加,甚至是在总体及预期生存时间减少的情况下。通过年龄、TNM分期分别对CS进行分层分析。年轻的患者、较低TNM分期,有更好的CS。结论与实际生存时间相比较,有关胰腺癌术后的预后评分系统低估了生存时间。条件生存率,在自然增加的寿命的基础上,比预期或实际生存时间都要好,这表明生存时间是动态变化的,而不是静止的。当把已经存活时间作为当前肿瘤未知生物学影响因素的一种标记时,CS也许能够更加合适的被用来预测生存时间。     

Enhanced high‐mobility group box 1 (HMGB1) modulates regulatory T cells (Treg)/T helper 17 (Th17) balance via toll‐like receptor (TLR)‐4‐interleukin (IL)‐6 pathway in patients with chronic hepatitis B

High‐mobility group box 1 (HMGB1) proteins are substantially up‐regulated in acute and chronic hepatitis. However, the immunopathogenic role of HMGB1 in patients with chronic hepatitis B (CHB) has not been elucidated. In this study, using a cohort of 36 CHB patients, we demonstrated a crucial role for HMGB1 to modulate balance between regulatory T (Treg) and T helper 17 (Th17) cells via the toll‐like receptor (TLR)‐4‐interleukin (IL)‐6 pathway. Serum HMGB1 levels were dramatically higher in CHB patients and increased along with liver injury, inflammation and fibrosis. Notably, HMGB1 increased along with decreased Treg/Th17 cells ratios in the periphery or intrahepatic microenvironment, which provides a clue for HMGB1 to favour Th17 responses whereas inhibit Treg responses. For in vitro studies, serum pools were constructed with serum from CHB patients at an advanced stage, whereas peripheral blood mononuclear cells (PBMC) pools were constructed with cells from those at an early stage. CHB‐serum significantly enhanced retinoic acid‐related orphan receptor‐γt (RORγt), whereas they inhibited forkhead box P3 (Foxp3) expression in CHB‐PBMC, which could be reversed by blocking of HMGB1, TLR4, or IL‐6. Besides, recombinant HMGB1 (rHMGB1) dose‐dependently up‐regulated RORγt whereas down‐regulated Foxp3 expression in CHB‐PBMC, and meanwhile, rHMGB1 enhanced TLR4 and IL‐6 expression in CHB‐PBMC. Moreover, the axis of HMGB1–TLR4‐IL‐6–Treg/Th17 required noncontact interactions between CD4 and non‐CD4 cells. In addition, rHMGB1 down‐regulated anti‐inflammatory proteins on CD4⁺CD25⁺ cells whereas up‐regulated pro‐inflammatory cytokines in CD4⁺CD25⁻ cells. In summary, enriched HMGB1 in CHB patients shifts Treg/Th17 balance to Th17 dominance via the TLR4‐IL‐6 pathway, which exacerbates liver injury and inflammation.     

Calcium Channel Blocker Compared With Angiotensin Receptor Blocker for Patients With Hypertension: A Meta‐Analysis of Randomized Controlled Trials

To explore the clinical effects of a calcium channel blocker compared with an angiotensin II receptor blocker in hypertensive patients, the authors collected data from randomized controlled trials. The pooled outcomes were all‐cause mortality, stroke, myocardial infarction, and heart failure. Eight head‐to‐head trials enrolling 25,084 patients were included. There was no significant mortality difference in the two arms (relative risk, 0.99; 95% confidence interval, 0.91–1.07). However, calcium channel blockers were more effective in reducing stroke (relative risk, 0.87; 95% confidence interval, 0.76–0.99) and myocardial infarction incidence (relative risk, 0.86; 95% confidence interval, 0.76–0.98). There was no significant difference with heart failure incidence between the two arms but a lower trend in patients with angiotensin II receptor blockers was noted (relative risk, 1.4; 95% confidence interval, 0.99–1.98). The meta‐analysis suggested that initially use of a calcium channel blocker might be superior to an angiotensin II receptor blocker for prevention of stroke and myocardial infarction.     

Engineering of an epoxide hydrolase for efficient bioresolution of bulky pharmaco substrates

Optically pure epoxides are essential chiral precursors for the production of (S)-propranolol, (S)-alprenolol, and other β-adrenergic receptor blocking drugs. Although the enzymatic production of these bulky epoxides has proven difficult, here we report a method to effectively improve the activity of BmEH, an epoxide hydrolase from Bacillus megaterium ECU1001 toward α-naphthyl glycidyl ether, the precursor of (S)-propranolol, by eliminating the steric hindrance near the potential product-release site. Using X-ray crystallography, mass spectrum, and molecular dynamics calculations, we have identified an active tunnel for substrate access and product release of this enzyme. The crystal structures revealed that there is an independent product-release site in BmEH that was not included in other reported epoxide hydrolase structures. By alanine scanning, two mutants, F128A and M145A, targeted to expand the potential product-release site displayed 42 and 25 times higher activities toward α-naphthyl glycidyl ether than the wild-type enzyme, respectively. These results show great promise for structure-based rational design in improving the catalytic efficiency of industrial enzymes for bulky substrates.Optically pure epoxides and the corresponding vicinal diols are valuable chiral building blocks for the production of pharmaceutically active compounds and other fine chemicals (1). Existing approaches for preparing enantiopure epoxides and diols include the asymmetric epoxidation or dihydroxylation of olefin substrates and the resolution of racemic epoxides. These reactions can be accomplished with either chemical catalysts such as chiral salen cobalt complexes and porphyrin manganese adducts or biocatalysts such as monooxygenases and epoxide hydrolases (EHs) (2–4). In the past two decades, EHs have received much attention because they are cofactor-independent enzymes that are “easy to use” for catalyzing the hydrolysis of racemic epoxides to yield highly enantiopure epoxides and vicinal diols (1, 5, 6). However, application of EHs in laboratory and industry was often hindered by their narrow substrate scope, low enantioselectivity, and regioselectivity, or product inhibition (7, 8).Many protein-engineering efforts have been made to overcome these drawbacks (9, 10). For example, directed evolution by error-prone PCR or DNA shuffling has been used to enhance the activity and enantioselectivity of EHs (11–13). Structure-guided mutagenesis also generated a few EH variants with improved catalytic performance (14–16). The strategy of iterative Combinatorial Active Site-Saturation Test (CAST) combines the rational approach and directed evolution to yield high-quality and small focused mutant libraries for screening EHs with better enantioselectivity (7, 17). By mutating residues at the substrate-binding site, the substrates of EHs have been expanded to include cyclic meso-epoxides, phenyl glycidyl ether (PGE) derivatives, and other styrene oxide-like analogs (18, 19). However, the catalytic efficiency of EH is still not satisfactory for bulky epoxide substrates including precursors of (S)-propranolol, (S)-alprenolol, and other β-adrenergic receptor blocking drugs (20, 21).In this work, we select BmEH, an EH cloned from Bacillus megaterium ECU1001, to expand its substrate scope for bulky pharmaco substrate α-naphthyl glycidyl ether (NGE). This enzyme is a potential industrial biocatalyst because it has unusual (R)-enantioselectivity and resolves ortho-substituted PGEs and para-nitrostyrene oxide with excellent enantiomeric ratios (E > 200) (22). We first identified the active tunnel of BmEH by solving its crystal structure complexed with a substrate analog phenoxyacetamide (POA) and analyzing the routes of substrate entry and product release by mass spectrum analysis. Alanine scanning experiments targeted to the potential product-release site of BmEH resulted in two variants, F128A and M145A, with efficient bioresolution abilities on NGE. Further kinetic measurements and structural analysis showed that M145A has much higher activity for the transition state intermediate formation, whereas both mutants exhibited expanded product-release site. The M145A BmEH variant has been successfully applied for the preparation of (S)-propranolol on a gram scale. The engineering of the potential product-release site described herein should have great promise for structure-based rational design of better industrial enzymes.     

Mechanistic roles for calcium and vitamin D in the regulation of body weight

Low intakes of calcium and inadequate vitamin D status often cluster with higher prevalence rates of obesity. Consequently, there has been much interest in the mechanisms by which calcium and vitamin D could regulate body weight and adiposity. This review has focused on randomized controlled trials (RCTs) that have manipulated these nutrients and studied pathways of energy balance. Overall, there is consistent evidence that calcium and vitamin D increase whole body fat oxidation after single and multiple meals, and that calcium promotes a modest energy loss through increased faecal fat excretion. The evidence is equivocal for a greater diet‐induced thermogenesis, increased lipolysis, suppression of key lipogenic enzymes, decreased hunger ratings or reduced energy/macronutrient intake. Emerging evidence suggests a potential improvement in insulin sensitivity following vitamin D that would impinge on food intake and substrate oxidation. However, the very few RCTs on supplemental vitamin D and energy balance have not explored postprandial avenues of the hormone's actions. Future efforts in this area need to define the threshold intake of these nutrients that would maximize metabolic and gastrointestinal outcomes. Such studies would provide a platform for endorsing the non‐skeletal role of calcium and vitamin D in human pathophysiology.     

高度近视合并脉络膜脱离型视网膜脱离预后相关因素分析

目的 分析高度近视合并脉络膜脱离型孔源性视网膜脱离（RRD-CD） 患者的预后相关因素。设计 回顾性病例系列。研究对象 2004-2018年北京同仁医院高度近视合并RRD-CD患者占836例。方法 回顾北京同仁医院住院HIS系统,收集高度近视合并RRD-CD手术治疗患者临床资料。随访6个月视网膜复位为复位组,发生视网膜再脱离为未复位组。采用Logistic回归法分析视网膜脱离复发的危险因素。主要指标 手术成功率及复发危险因素。结果 共纳入高度近视合并RRD-CD患者836例,平均年龄（56.51±12.14）岁;男性518例（61.9%）,右眼发病434例 （51.9%）。视网膜脱离未复位为22.7%,与视网膜复位患者相比,未复位患者年龄较轻,术前视力、眼轴、晶状体状态、视网膜裂孔、增生性玻璃体视网膜病变（PVR）等级及手术方式等差异均有统计学意义,其中年龄〔优势比（OR）=0.972,95%可信区间（95%CI）,0.967~0.989〕,术前视力光感（OR=1.898,95%CI为1.297~2.777）,人工晶状体眼（OR=1.860,95%CI为1.255~2.758）,眼轴>30 mm（OR=1.718,95%CI为1.240~2.379）,巨大视网膜裂孔（OR=2.464,95%CI为1.495~4.063）及PVR D级（OR=1.551,95%CI为1.046~2.300）为视网膜脱离未复位的危险因素。结论 高度近视合并RRD-CD患者男性比例大,中年发病、超高度近视、人工晶状体眼、巨大视网膜裂孔及PVD D级患者首次手术成功率低,视网膜脱离易复发。（眼科,2021,30： 42-46）     

Peripheral arterial filling time and peripheral retina fluorescence features in ultra-widefield angiography

AIM: To evaluate the peripheral arterial filling time (PAFT) and venous filling time (VFT) in eyes without known diseases that may influence filling process using ultra-widefield (UWF) fluorescein angiography (FA), and to review the peripheral retina fluorescence features. METHODS: A total of 30 eyes of 30 patients were retrospectively reviewed in this observational study. UWF-FA was performed using Optos 200Tx. PAFT and VFT was recorded. The interval between the arterial or venous filling completion and the previous photo was documented. The appearance of the far peripheral retina was described as either granular background fluorescence or mottled fluorescent band or vascular leakage. Terminal vascular patterns was described as loop pattern or branching pattern. Microvascular abnormalities such as arteriovenous shunting, vessels crossing the horizontal raphe, right angle vessels, terminal networks, capillary nonperfusion, drusen or microaneurysms were evaluated. RESULTS: The normal limits of PAFT was 3.397-8.984s and 4.399-11.753s for VFT. The appearance of the far peripheral retina, defined as granular background (63%), mottled fluorescence (20%), or vascular leakage (17%), was symmetrical between both eyes. Capillary nonperfusion (23%) and microaneurysms (40%) were more frequently found in eyes with loop pattern than in eyes with branching pattern. Other peripheral signs such as right angle vessels (73%), and terminal networks (80%) were commonly seen on UWF-FA in the normal peripheral retina. CONCLUSION: The main courses of retinal artery and vein filling time are overlapping with each other on UWF-FA. Notably, the arterial filling process is completed in the arteriovenous phase rather than the traditionally named arterial phase. There are various manifestations in the peripheral retina of normal eyes.