Homeostatic regulation of T follicular helper and antibody response to particle antigens by IL-1Ra of medullary sinus macrophage origin

Hepatitis B virus (HBV) vaccines are composed of surface antigen HBsAg that spontaneously assembles into subviral particles. Factors that impede its humoral immunity in 5% to 10% of vaccinees remain elusive. Here, we showed that the low-level interleukin-1 receptor antagonist (IL-1Ra) can predict antibody protection both in mice and humans. Mechanistically, murine IL-1Ra–inhibited T follicular helper (Tfh) cell expansion and subsequent germinal center (GC)-dependent humoral immunity, resulting in significantly weakened protection against the HBV challenge. Compared to soluble antigens, HBsAg particle antigen displayed a unique capture/uptake and innate immune activation, including IL-1Ra expression, preferably of medullary sinus macrophages. In humans, a unique polymorphism in the RelA/p65 binding site of IL-1Ra enhancer associated IL-1Ra levels with ethnicity-dependent vaccination outcome. Therefore, the differential IL-1Ra response to particle antigens probably creates a suppressive milieu for Tfh/GC development, and neutralization of IL-1Ra would resurrect antibody response in HBV vaccine nonresponders.

Follicular helper T (Tfh) cells are antigen-experienced CD4⁺ T cells within B cell follicles of secondary lymphoid organs, such as lymph nodes (LN), spleens, and Peyer’s patches, that constitutively express the B cell follicle homing receptor CXCR5 (1). Upon cellular interaction and cross-signaling with their cognate follicular B (FoB) cells in the presence of follicular dendritic cells (FDCs), Tfh cells trigger the formation and maintenance of germinal centers (GCs) through the expression of CD40 ligand and the secretion of IL-21 and IL-4 (2–4). Tfh-dependent paracrine activation of CD40 results in B cell survival and differentiation in the GC (5), whereas isotype class switching is believed to occur predominantly outside GCs. Therefore, Tfh cells play a critical role in mediating the selection of high-affinity B cells that differentiate either into plasma cells or memory B cells (6–11).Besides the inducible T cell costimulator (ICOS) that activates Tfh cells to secrete IL-21, other cytokines [e.g., IL-2 (12), IL-6 (13), and IL-7 (14)] also signal for Tfh cell differentiation. The role of IL-1 signaling remained puzzling until recently: Tfh cells can be primed by IL-1β, whose production is licensed by IFN-β in response to infectious agents (15). Such featured innate response of IFN-β and IL-1β relies on the activation of TLR and inflammasomes by live, but not dead, bacteria or recombinant vaccines (16, 17). Therefore, OVA antigen augments Tfh cell response in mice only when IL-1β is exogenously applied at a nonphysiological high concentration (18), whereas endogenous IL-1β/IL-1R1 signaling may not be required for antibody responses to T-dependent or -independent antigens (19–21). We reasoned that IL-1Ra (encoded by IL-1rn), which can compete with IL-1 for binding to IL-1R1 in the homeostatic inflammatory response (22–24), would intrinsically antagonize IL-1β/IL-1R1 signaling for Tfh/GC development. For example, IL-1rn^−/− mice exhibit an excessive antibody response to sheep red blood cells immunization (25, 26). A thorough investigation is required to dissect how IL-1 and IL-1Ra mutually regulate a homeostatic Tfh/GC response.LN macrophages are conventionally divided into two subtypes. Subcapsular sinus (SCS, CD169⁺F4/80⁻) macrophages are specialized antigen presenting cells that capture certain particle or opsonized antigens and display them intact for cognate recognition by FoB cells (27–30). SCS macrophages also relay immune complex to noncognate B cells for antibody affinity maturation (30). Macrophages in medullary sinus (MSM, CD169⁺F4/80⁺), in contrast, are potent in phagocytosis (31) for clearance of pathogens and particulate antigens from the lymph. It has been postulated 10 y ago that SCS may capture particle antigens, such as hepatitis B virus (HBV) vaccine, and migrate to follicles to facilitate more effective activation of B cells and FDCs (32). In this work, we found that murine antibody response inversely correlated to IL-1Ra level and clearly distinguished responders from nonresponders in volunteers receiving HBV vaccination. Further studies showed that LN macrophages subsets exhibited different capture and activation kinetics for particle and soluble antigens, and IL-1Ra expression by MSM could critically modulate IL-1R1 potentiation of Tfh cells and, hence, the specific antibody response to particle antigens. Therefore, mice lacking IL-1Ra or with IL-1Ra being neutralized yielded more robust antibody response to HBV vaccine and enables protection against chronic HBV infection.     

紫雪丹的质量控制

目的:建立紫雪丹的质量控制标准。方法:采用薄层色谱法对甘草、木香进行定性鉴别;采用气相色谱法对龙脑进行含量测定。结果:定性鉴别阴性无干扰,分离度高;龙脑在100μg·m L-1~800μg·m L-1(r=0.9996)范围内线性关系良好,加样回收率为100.79%,RSD为2.5%(n=6)。结论:方法可用于紫雪丹的质量控制。     

酚氨咖敏颗粒质量标准研究

目的完善酚氨咖敏颗粒的质量标准。方法采用薄层色谱(TLC)法对乙酰氨基酚、氨基比林、咖啡因及马来酸氯苯那敏进行定性鉴别。采用气相色谱(GC)法同时测定4种成分的含量,色谱柱为TR-1弹性石英毛细管柱(30 m×0. 32 mm,0. 25μm),程序升温(起始温度为180℃,保持1 min,再以8℃/min的速率升至240℃,保持2 min),氢火焰离子化检测器温度为260℃。结果 TLC法可同时鉴别酚氨咖敏颗粒中4种成分。对乙酰氨基酚、氨基比林、咖啡因及马来酸氯苯那敏质量浓度线性范围分别为186. 3~5 960μg/m L(r=0. 999 9),124. 9~3 996μg/m L(r=0. 999 9),38. 24~1 224μg/m L(r=1. 000 0)和2. 525~80. 80μg/m L(r=0. 999 9);平均加样回收率分别为98. 69%,99. 45%,99. 52%,101. 40%,RSD分别为1. 36%,1. 62%,1. 20%,1. 25%(n=9)。结论该方法简便快速、准确可靠,适用于酚氨咖敏颗粒中4种成分的定性、定量分析,可更全面地控制该制剂的质量。     

Analysis of methanol and its derivatives in illegally produced alcoholic beverages

IntroductionIllegal alcohol production remains as a common issue worldwide. Methanol poisoning mostly occurs because of the methanol used in production of counterfeit alcohol instead of ethyl alcohol due to its low price or by drinking the liquids containing methyl alcohol. Pectolytic enzymes results in an increase of methanol levels in many fermentation products such as ciders or wines. Methanol poisonings are infrequently encountered in forensic medicine practice. However, sporadic cases due to methanol intoxication as well as epidemic cases have been reported. In this study, we aimed to identify existence of methanol and its metabolites in illegally produced alcoholic beverages used in Antakya region.Material and methodsTwelve legally produced alcohol samples and Fifty-six different illegally produced alcohol samples were collected from the markets and local producers. Existence of methanol, formic acid, methyl amine, methyl formate and trioxan were determined using GC–MS method in these samples.ResultsFifty-six different illegal alcohol samples were analyzed in this study and methanol was detected in 39 (75%) of samples. Formic acid was detected in 3, formamide in 1, methyl amine in 6, methyl formate in 10 and trioxan in 2 samples.ConclusionOverwhelming majority of illegal alcoholic beverages was detected to contain methanol. Interestingly this study also revealed the presence of trioxane, which has not previously reported among toxic agents in illegal alcohol samples.     

Metabolic changes in rats after intragastric administration of MGCD0103 (Mocetinostat), a HDAC class I inhibitor

MGCD0103, an isotype-selective HDACi, has been clinically evaluated for the treatment of hematologic malignancies and advanced solid tumors, alone and in combination with standard-of-care agents. In this study, we developed a serum metabolomic method based on gas chromatography-mass spectrometry (GC-MS) to evaluate the effect of intragastric administration of MGCD0103 on rats. The MGCD0103 group rats were given 20, 40, 80 mg/kg of MGCD0103 by intragastric administration each day for 7 days. Pattern recognition analysis, including both principal component analysis (PCA) and partial least squares-discriminate analysis (PLS-DA) revealed that intragastric administration of MGCD0103 induced metabolic perturbations. As compared to the control group, the levels of L-alanine, L-isoleucine, and L-leucine of MGCD0103 group decreased. The results indicate that metabolomic methods based on GC-MS may be useful to elucidate side effect of MGCD0103 through the exploration of biomarkers (L-alanine, L-isoleucine, and L-leucine). According to the pathological changes of liver at difference dosage, MGCD0103 is hepatotoxic and its toxity is dose-dependent.     

Sex differences in circadian timing systems: Implications for disease

Virtually every eukaryotic cell has an endogenous circadian clock and a biological sex. These cell-based clocks have been conceptualized as oscillators whose phase can be reset by internal signals such as hormones, and external cues such as light. The present review highlights the inter-relationship between circadian clocks and sex differences. In mammals, the suprachiasmatic nucleus (SCN) serves as a master clock synchronizing the phase of clocks throughout the body. Gonadal steroid receptors are expressed in almost every site that receives direct SCN input. Here we review sex differences in the circadian timing system in the hypothalamic–pituitary–gonadal axis (HPG), the hypothalamic–adrenal–pituitary (HPA) axis, and sleep–arousal systems. We also point to ways in which disruption of circadian rhythms within these systems differs in the sexes and is associated with dysfunction and disease. Understanding sex differentiated circadian timing systems can lead to improved treatment strategies for these conditions.     

GC法测定一次性肠外营养输液袋中乙酸乙烯酯迁移量

目的:建立GC法测定一次性肠外营养输液袋中乙酸乙烯酯的迁移量.方法:以DB-624毛细管色谱柱为固定液;程序升温:起始温度90℃,维持5 min,以每分钟50℃的速率升温至250℃,维持5 min;进样口温度为220℃;检测器温度为250℃;流速为1.0 mL·min-1;分流比为10:1.结果:本法可将溶剂、空白溶液与乙酸乙烯酯单体较好分离,乙酸乙烯酯在4.47~10.44μg·mL-1(r=0.9993)浓度范围内线性关系良好,回收率为100.4%,RSD为1.25%.结论:本法灵敏度高,操作简单、快速、准确,适合输液袋中乙酸乙烯酯单体迁移量的监测.     

气相色谱法测定复方樟脑搽剂中樟脑与薄荷脑含量及其不确定度评估

目的 建立测定复方樟脑搽剂中主要成分樟脑和薄荷脑含量的气相色谱（GC）法,并探讨不确定度评估方法。方法 采用GC法,以萘为内标物,Thermo TG-WAXMS毛细管柱（30.0 m×0.25 mm,涂层厚0.5 μm）,FID检测器;分析测量过程不确定度的来源,建立数学模型,计算合成标准不确定度和扩展不确定度。结果 樟脑和薄荷脑分别在8.56~85.6μg/ml、11.2~112.0 μg/ml质量浓度范围内线性关系良好,樟脑平均加样回收率100.20%、RSD为0.892%（n=9）,薄荷脑平均加样回收率99.65%、RSD为1.369%（n=9）;扩展不确定度分别为0.630 mg/ml （k=2）、0.656 mg/ml （k=2）,樟脑含量为（20.74±0.630） mg/ml、薄荷脑含量为（20.48±0.656） mg/ml。结论 气相色谱方法准确、灵敏、重现性好,可用于复方樟脑搽剂中樟脑和薄荷脑含量测定;不确定度评定有利于进一步提高含量测定的准确性。     

GC测定非诺贝特制剂中十二烷基硫酸钠含量

目的 建立非诺贝特制剂中十二烷基硫酸钠的GC测定方法。方法 采用HP-5（30 m×0.53 mm,2.65 μm）毛细管柱,以氮气为载气,氢火焰离子化检测器,程序升温,直接进样,以十二烷醇为对照品外标法测定十二烷基硫酸钠的含量。结果 十二烷醇在0.165~1.32 mg·mL^-1内具有良好的线性关系,相关系数为0.999 5。国内企业产品的十二烷基硫酸钠用量远低于安全用量,但部分国内企业可能存在过量添加和非法添加的问题。结论 本方法准确,专属性强,可作为非诺贝特制剂中十二烷基硫酸钠的检测方法。