Targeting proinsulin to local immune cells using an intradermal microneedle delivery system; a potential antigen-specific immunotherapy for type 1 diabetes.
使用皮内微针递送系统将胰岛素原靶向于局部免疫细胞; 1 型糖尿病的潜在抗原特异性免疫治疗。
- 作者列表："Arikat F","Hanna SJ","Singh RK","Vilela L","Wong FS","Dayan CM","Coulman SA","Birchall JC
:Antigen-specific immunotherapy (ASI) has been proposed as an alternative treatment strategy for type 1 diabetes (T1D). ASI aims to induce a regulatory, rather than stimulatory, immune response in order to reduce, or prevent, autoimmune mediated β-cell destruction, thus preserving endogenous insulin production. The abundance of immunocompetent antigen presenting cells (APCs) within the skin makes this organ an attractive target for immunotherapies. Microneedles (MNs) have been proposed as a suitable drug delivery system to facilitate intradermal delivery of autoantigens in a minimally invasive manner. However, studies to date have employed single peptide autoantigens, which would restrict ASI to patients expressing specific Human Leukocyte Antigen (HLA) molecules, thus stratifying the patient population. This study aims to develop, for the first time, an intradermal MN delivery system to target proinsulin, a large multi-epitope protein capable of inducing tolerance in a heterogenous (in terms of HLA status) population of T1D patients, to the immunocompetent cells of the skin. An optimized three component coating formulation containing proinsulin, a diluent and a surfactant, facilitated uniform and reproducible coating of >30 μg of the active pharmaceutical ingredient on a stainless steel MN array consisting of thirty 500 μm projections. When applied to a murine model these proinsulin-coated MNs efficiently punctured the skin and after a limited insertion time (150 s) a significant proportion of the therapeutic payload (86%) was reproducibly delivered into the local tissue. Localized delivery of proinsulin in non-obese diabetic (NOD) mice using the coated MN system stimulated significantly greater proliferation of adoptively transferred antigen-specific CD8+ T cells in the skin draining lymph nodes compared to a conventional intradermal injection. This provides evidence of targeted delivery of the multi-epitope proinsulin antigen to skin-resident APCs, in vivo, in a form that enables antigen presentation to antigen-specific T cells in the local lymph nodes. The development of an innovative coated MN system for highly targeted and reproducible delivery of proinsulin to local immune cells warrants further evaluation to determine translation to a tolerogenic clinical outcome.
: 抗原特异性免疫治疗 (ASI) 已被提出作为 1 型糖尿病 (T1D) 的替代治疗策略。ASI 旨在诱导一种调节性的，而不是刺激性的免疫反应，以减少或防止自身免疫介导的 β 细胞破坏，从而保留内源性胰岛素的产生。皮肤内丰富的免疫活性抗原呈递细胞 (apc) 使该器官成为免疫疗法的一个有吸引力的靶点。微针 (MNs) 被认为是一种合适的药物递送系统，以微创方式促进自体抗原的皮内递送。然而，迄今为止的研究采用了单肽自身抗原，这将限制 ASI 患者表达特定的人类白细胞抗原 (HLA) 分子，从而对患者人群进行分层。本研究旨在首次开发一种皮内 MN 递送系统，靶向胰岛素原，一种能够诱导异源 (HLA 状态) 耐受的大型多表位蛋白 t1D 患者群体，皮肤的免疫活性细胞。包含胰岛素原、稀释剂和表面活性剂的优化的三组分涂层配方, 在由 30 个 500 μ m 投影组成的不锈钢 MN 阵列上促进了> 30 μ g 活性药物成分的均匀和可重复涂层。当应用于小鼠模型时，这些胰岛素原包被的 MNs 有效地刺穿皮肤，并且在有限的插入时间 (150 s) 后，相当大比例的治疗有效载荷 (86%) 可重复递送到局部组织中。非肥胖糖尿病患者胰岛素原的局部递送 (NOD) 与常规皮内注射相比，使用包被 MN 系统的小鼠刺激了皮肤引流淋巴结中过继转移的抗原特异性 CD8 + T 细胞的显著更大增殖。这提供了多表位胰岛素原抗原靶向递送至皮肤驻留的 apc 的证据，体内, 以使抗原呈递到局部淋巴结中的抗原特异性 T 细胞的形式。开发一种创新的包被 MN 系统，用于将胰岛素原高度靶向和可重复递送到局部免疫细胞，值得进一步评价以确定向耐受性临床结果的转化。
METHODS:BACKGROUND:Given the importance of habitual dietary protein intake, distribution patterns and dietary sources in the aetiology of age-related declines of muscle mass and function, the present study examined these factors as a function of sex and age in Irish adults aged 18-90 years comprising The National Adult Nutrition Survey (NANS). METHODS:In total, 1051 (males, n = 523; females, n = 528) undertook a 4-day semi-weighed food diary. Total, body mass relative intake and percentage contribution to total energy intake of dietary protein were determined in addition to protein distribution scores (PDS), as well as the contribution of food groups, animal- and plant-based foods to total protein intake. RESULTS:Total and relative protein intake [mean (SD)] were highest in those aged 18-35 years [96 (3) g day , 1.32 (0.40) g kg day ], with lower protein intakes with increasing age (i.e. in adults aged ≥65 years [82 (22) g, 1.15 (0.34) g kg day , P < 0.001 for both]. Differences in protein intake between age groups were more pronounced in males compared to females. Protein distribution followed a skewed pattern for all age groups [breakfast, 15 (10) g; lunch, 30 (15) g; dinner, 44 (17) g]. Animal-based foods were the dominant protein source within the diet [63% (11%) versus 37% (11%) plant protein, P < 0.001]. CONCLUSIONS:Protein intake and the number of meals reaching the purported threshold for maximising post-prandial anabolism were highest in young adults, and lower with increasing age. For main meals, breakfast provided the lowest quantity of protein across all age categories and may represent an opportunity for improving protein distribution, whereas, in older adults, increasing the number of meals reaching the anabolic threshold regardless of distribution pattern may be more appropriate.
METHODS:BACKGROUND:Low cardiorespiratory fitness (CRF) increases risk of all-cause mortality and cardiovascular events. Periodic CRF assessment can have an important preventive function. OBJECTIVE:To develop a protocol-free method to estimate CRF in daily life based on heart rate (HR) and body acceleration measurements. METHODS:Acceleration and HR data were collected from 37 subjects (M=49%) while performing a standardized laboratory activity protocol (sitting, walking, running, cycling) and during a 5-days free-living monitoring period. CRF was determined by oxygen uptake (VO2max) during maximal exercise testing. A doubly-labeled water validated equation was used to predict total energy expenditure (TEE) from acceleration data. A fitness index was defined as the ratio between TEE and HR (TEE-pulse). Activity recognition techniques were used to process acceleration features and classify sedentary, ambulatory and other activity types. Regression equations based on TEE-pulse data from each activity type were developed to predict VO2max. RESULTS:TEE-pulse measured within each activity type of the laboratory protocol was highly correlated to VO2max (r from 0.74 to 0.91). Averaging the outcome of each activity-type specific equation based on TEE-pulse from the laboratory data led to accurate estimates of VO2max (RMSE: 300.0 mlO2/min or 10%). The difference between laboratory and free-living determined TEE-pulse was 3.7 ± 11% (r =0.85). The prediction method preserved the prediction accuracy when applied to free-living data (RMSE: 367 mlO2/min or 12%). CONCLUSIONS:Measurements of body acceleration and HR can be used to predict VO2max in daily life. Activity-specific prediction equations are needed to achieve highly accurate estimates of CRF.
METHODS:OBJECTIVE:Postprandial dyslipidemia is a common feature of insulin resistant states and contributes to increased cardiovascular disease risk. Recently, bile acids have been recognized beyond their emulsification properties as important signaling molecules that promote energy expenditure, improve insulin sensitivity, and lower fasting lipemia. While bile acid receptors have become novel pharmaceutical targets, their effects on postprandial lipid metabolism remain unclear. Here we investigated the potential role of bile acids in regulation of postprandial chylomicron production and triglyceride excursion. Approach and Results: Healthy C57BL/6 mice were given an intraduodenal infusion of taurocholic acid (TA) under fat-loaded conditions and circulating lipids were measured. Targeting of bile acid receptors was achieved with GW4064, a synthetic agonist to the farnesoid X receptor (FXR), and with deoxycholic acid (DCA), an activator of the Takeda G-protein-coupled receptor 5. TA, GW4064, and DCA treatments all lowered postprandial lipemia. FXR agonism also reduced intestinal triglyceride content and activity of microsomal triglyceride transfer protein, involved in chylomicron assembly. Importantly, TA effects (but not DCA) were largely lost in FXR knockout mice. These bile acid effects are reminiscent of the anti-diabetic hormone glucagon-like peptide-1 (GLP-1). While the GLP-1 receptor agonist exendin-4 retained its ability to acutely lower postprandial lipemia during bile acid sequestration and FXR deficiency, it did raise hepatic expression of the rate limiting enzyme for bile acid synthesis. CONCLUSIONS:Bile acid signaling may be an important mechanism of controlling dietary lipid absorption and bile acid receptors may constitute novel targets for the treatment of postprandial dyslipidemia.