慢性乙型肝炎患者树突状细胞的培养鉴定和功能特点

目的：研究慢性乙型肝炎患者树突状细胞（DC）的功能特点,探讨其与肝炎发病机制的关系。方法：分离获得18例慢性乙型肝炎患者和10例正常人外周血单个核细胞（PBMC）,在体外培养条件下,加入细胞因子重组人粒细胞巨噬细胞集落刺激因子（GM－CSF）,IL－4,胎肝酪氨酸激酶受体的配体（FLt3-L)和肿瘤坏死因子α（TNFα）,使DC细胞增殖,成熟,用流式细胞仪检测DC的表面标志,同时检测DC在混合淋巴细胞反应（MLR）中的刺激能力和细胞因子的分泌。结果：DC在体外经细胞因子的刺激可明显增殖,但慢性乙型肝炎患者DC的增殖速度低于正常人;DC表面标志人组织相容性复合物Ⅱ类分子（HLA－DR）,CD－80（B7－1）,CD－86（B7－2）和CD－1α的表达较正常对照组均明显降低（P＜0．001）,尤以CD－1α的降低更为显著,DC在MLR中的刺激能力亦明显低于正常对照。并且与正常对照相比,其产生的IL－12水平降低,而NO水平却增高（P＜0．05）。结论：慢性乙型肝炎患者DC的表型不成熟和功能的缺失,由此导致IL－12的产生和刺激T细胞增殖能力的降低,可能是HBV感染持续发展的原因之一。     

补肾健脾解毒法中药复方对慢性HBV携带者树突状细胞的影响

目的：研究补肾健脾解毒法中药复方对慢性HBV携带者树突状细胞（DC）的影响。方法：研究分为两个试验组。治疗组纳入30例慢性HBV携带者,对照组纳入10例健康志愿者,分别采用补肾健脾解毒法中药复方、安慰剂干预48周,观察两组人员治疗前后DC的成熟表面标志物、同种混合淋巴细胞反应（MLR）及其细胞因子（IL-12、IFN-γ）的变化。结果：基线时,治疗组患者的DC成熟表面标记物HLA（人白细胞抗原）-DR、CD86、CD80和CD1α的表达水平分别为61.34%、55.78%、42.63%、43.17%,明显低于对照组的90.52%、83.26%、76.81%、84.90%（P〈0.01）;DC诱导MLR的能力、MLR上清液中IL-12和IFN-γ的表达水平全都明显低于对照组（P〈0.01）。治疗48周后,治疗组DC成熟表面标记物的表达水平均较治疗前明显上升（P〈0.01）,但仍显著低于对照组（P〈0.01）;治疗组患者对DC诱导MLR能力也有显著提升,MLR上清液中IL-12和IFN-γ的表达水平明显上升（P〈0.01）,但DC诱导MLR能力及IFN-γ的表达水平未达到对照组水平（P〈0.05或0.01）。结论：补肾健脾解毒法中药复方能促进慢性HBV携带者的DC成熟并改善其免疫功能,从而产生有效的免疫应答以抑制/清除HBV。     

乙型肝炎相关肝癌外周血树突状细胞负载肿瘤抗原前后免疫功能的变化

目的探讨肝癌患者外周血单个核细胞(PBMC)来源树突状细胞(DC)表面分子表达及负载肿瘤抗原前后免疫功能变化与免疫逃逸的关系。方法分离18例乙型肝炎相关原发性肝癌、11例乙型肝炎肝硬化患者和10名健康献血者PBMC,体外培养,并加入重组人粒细胞巨噬细胞集落刺激因子(GM-CSF)和白细胞介素-4(IL-4)诱导DC。以共聚焦显微镜和扫描电镜观察形态,以流式细胞仪检测DC表面人类白细胞抗原(HLA)-DR、CD1a、CD80、CD83、CD86等分子表达水平。以HCCLM6肝癌细胞株制备肿瘤抗原,分别负载3种DC,最后以混合淋巴细胞反应(MLR)测定DC负载前后刺激同种异型T淋巴细胞增殖能力,并测定MLR上清液中IL-12的含量。结果肝硬化和肝癌组PBMC、DC得率低于正常组(P<0.05);HLA- DR、CD1a、CD80和CD86等分子表达水平也低于正常组(P<0.05);负载肿瘤抗原前肝硬化和肝癌组刺激同种异型T淋巴细胞增殖能力和MLR上清液中IL-12含量明显低于正常组,负载肿瘤抗原后3组均提高, 并以肝硬化组提高最为明显,但IL-12含量仍低于正常组。结论DC表型和功能缺陷可能是乙型肝炎病毒产生免疫耐受和肝癌细胞免疫逃逸的重要机制。肝硬化患者DC仍有一定功能。     

含CpG基序寡核苷酸对慢性乙型肝炎患者外周血树突状细胞的免疫激活研究

目的 研究含非甲基化CpG基序的寡核苷酸（CpG-ODN）对慢性乙型肝炎患者（CHB）外周血树突状细胞（DC）表型和功能的影响。方法以CD14磁性分选微珠分离CHB患者外周血高纯度单核细胞；以重组人粒细胞巨噬细胞集落刺激因子（hGM—CSF）、重组人白细胞介素-4（hIL-4）诱导扩增DC；以CpG-ODN刺激DC成熟，并与肿瘤坏死因子（TNF）-n比较，评价其对DC表达表面分子人类白细胞组织相容性抗原（HLA）-DR、CD86、CD1a，分泌IL-12p70以及刺激同种T细胞增殖能力的影响。结果与non—ODN和磷酸盐缓冲液（PBS）组比较，CpG-ODN能明显提高CHB患者外周血DC表面分子HLA—DR、CD86的表达，使IL-12分泌增加，刺激同种异体T细胞增殖的能力亦显著增强（P=0．017和0．023），但不能明显提高CD1a的表达；CpG-ODN的上述刺激作用接近或略逊于hTNF-α，但差异无统计学意义（P〉0．05）。结论CpG-ODN与hTNF-α一样能够促进CHB患者外周血DC成熟进而增强其抗原提呈功能。     

慢性乙型肝炎患者外周血树突状细胞功能的研究

目的研究慢性乙型肝炎患者外周血树突状细胞(DC)的免疫功能.方法从慢性乙型肝炎患者外周血中分离单个核细胞,用无血清培养法分离培养DC,用流式细胞仪检测DC表型,用液闪计数仪观察DC对T细胞的增殖作用,用ELISA法检测混合淋巴细胞反应(MLR)中细胞因子的分泌水平.结果患者组DC的CD86的表达率为(70.2±5.2)%,明显低于正常人组(95.3±3.8)%,P＜0.01;其诱导T细胞增殖能力每分钟液闪计数cpm为10000±2000,明显低于正常人组(cpm为30000±3000),P＜0.01患者MLR中IL-12为(120.0±19.7)pg/ml,γ-干扰素为(799.0±161.3)pg/m1,明显低于正常人组的(280.0±41.1)pg/ml和(3359.0±635.4)pg/ml,P＜0.01.结论慢性乙型肝炎患者外周血DC免疫功能低下,并与DC表面CD86的表达率下降及DC分泌IL-12减少密切相关.     

无血清培养液培养大肠癌病人外周血来源树突状细胞

目的使用无血清培养液培养大肠癌病人外周血来源树突状细胞（DC）。方法采集大肠癌病人外周血分离出单核细袍。加入分重组人白细胞介素-4和重组人粒-巨噬细胞集落刺激因子的无血清培养液进行体外诱导培养，培养7d后获得DC。显微镜下观察DC形态，流式细胞仪检测细胞表型CD1a，CD14，CD80，CD83，CD86和HLA—DR的表达。结果无血清培养液培养的细胞呈毛刺状，流式细胞仪检测细胞表达CD1a、CD80、CD86和HLA-DR，不表达CD14、CD83，符合DC的典型特征。结论无血清培养液可以从大肠癌病人外周血诱导培养出DC。     

慢性乙型肝炎患者外周血树突状细胞的成熟度研究

目的：研究慢性乙型肝炎（CHB）患者外周血树突状细胞（DC）的成熟度及其占单核细胞的比例,探讨CHB患者DC免疫治疗的机制。方法选取20例CHB患者和10例健康人,分别采集外周抗凝全血2 mL,流式细胞仪检测DC表面CD80、CD86、CD83的表达量及单核细胞表面CD14的表达。结果两组DC表面CD80、CD86、CD83分子表达无差异（P＞0．05）;CHB组较健康对照组单核细胞表面分子 CD14表达明显增多,具显著统计学差异（P＜0．05）;CHB组CD80、CD86、CD83三者表达之和占CD14的比例明显高于CHB组,两者有显著统计学意义（P＜0．05）。结论CHB患者外周血存在单核细胞增多、成熟DC减少现象,恢复DC功能是治疗CHB的手段之一。     

PvMSP1对树突状细胞分化成熟和功能的影响

目的 目的 观察间日疟原虫裂殖子主要蛋白1 （PvMSP1） 对树突状细胞 （DC） 分化成熟和功能的影响, 并探讨该蛋白通 过Toll样受体 （TLR） 通路活化DC的机制。方法 方法 选择不同剂量的PvMSP1 （1.0、 10.0、 100.0 μg/ml） 体外刺激人单核细胞来 源的DC, 采用流式细胞术分析DC成熟性相关分子CD83、 CD86、 HLA?DR的表达变化; ELISA检测DC培养上清中IL?10、 IL?12 的表达水平; RT?PCR检测DC TLR4、 TLR9 mRNA的表达水平; MTT法检测DC刺激自体淋巴细胞增殖的能力。同时选择未 刺激的DC作为阴性对照组, LPS刺激的DC作为阳性对照组。对所得数据进行方差分析和q检验。结果 结果 与未刺激组比 较, LPS诱导组CD83、 CD86、 HLA?DR的百分含量均增加, PvMSP1诱导组CD83、 CD86、 HLA?DR的表达也均升高 （P均 < 0.05）; LPS诱导组IL?10、 IL?12的表达量明显增加 （P < 0.01）, PvMSP1诱导组IL?10、 IL?12的表达量也均增加 （P均 < 0.05）; LPS组DC TLR4 mRNA的表达增加 （P < 0.05）, TLR9 mRNA的表达无明显变化 （P > 0.05）, PvMSP1诱导组DC TLR4 mRNA 的表达增加 （P < 0.01）, TLR9 mRNA无明显变化 （P > 0.05）; DC能够刺激自体淋巴细胞增殖。结论 结论 PvMSP1具有促进DC 分化成熟的作用, 且经其诱导成熟的DC具备抗原递呈功能; PvMSP1可能经TLR4通路而非TLR9通路诱导DC成熟。     

HBsAg、HBcAg活化树突状细胞治疗慢性乙型肝炎的体外研究

目的 研究慢性乙型肝炎患者外周血树突状细胞(DC)经HBsAg、HBcAg活化后的免疫功能.方法 从慢性乙型肝炎患者外周血中培养扩增DC,在DC成熟前,加入纯的HBsAg、HBcAg刺激,用流式细胞仪检测DC表型,用液闪计数仪观察DC对T细胞的增殖作用,用ELISA法检测混合淋巴细胞反应(MLR)中细胞因子的分泌水平.结果 经HBcAg刺激DC的CD86表达率为(92.14±5.12)%,明显高于HBsAg刺激组和未加抗原组(P＜0.01);经HBcAg刺激组DC诱导同种异体静止T细胞增殖的能力每分钟液闪计数值(cpm)为34259±3127,明显高于HBsAg刺激组(20258±2917)和单个核细胞组(3469±417),P＜0.01;经HBcAg刺激组DC MLR中IL-12浓度为(342±42.3)ng/L,分别高于HBsAg刺激组和未加抗原组(P＜0.01).结论 体外经HBcAg刺激DC可有效提呈抗原病毒,并可进一步刺激T细胞产生.     

中药辨证干预对慢性乙型肝炎肾虚患者树突状细胞功能影响

目的研究中药辨证干预对慢性乙型肝炎（CHB）肾虚患者外周血树突状细胞（DC）功能的影响。方法 CHB肾虚证患者60例,健康对照组10例,分离各组外周血单个核细胞,体外诱导培养DC,流式细胞仪检测DC表面分子CD80、CD86、CD1a、HLA-DR表达率;酶联免疫吸附检测（ELISA）检测DC培养上清液中细胞因子IL-10表达水平,同时检测CHB患者血清HBV DNA定量并记录中医证候积分。予中药辨证干预治疗3个月,检测患者治疗后外周血DC表面分子CD80、CD86、CD1a、HLA-DR及培养上清中IL-10的表达水平及血清HBV DNA定量,并记录中医证候积分的变化。结果治疗前CHB肾虚证患者CD80、CD86、CD1a、HLA-DR表达率显著低于正常对照组（P〈0.05）,IL-10表达水平显著高于正常对照组（P〈0.05）;治疗后CHB肾虚证患者CD80、CD86、CD1a、HLA-DR表达率明显提高、IL-10表达水平明显降低（P〈0.05）,中医证候积分明显改善（P〈0.05）。结论 CHB肾虚证患者存在DC成熟障碍及功能异常,主要表现为表面共刺激分子表达率下降,抑制因子表达率升高。中药辨证干预不仅能改善患者中医证候积分,同时能提高DC表面共刺激分子表达率,降低抑制因子表达率,提示DC可能与中医虚实病机存在一定相关性。     

The immunoneuroendocrine role of melatonin

Abstract: A tight, physiological link between the pineal gland and the immune system is emerging from a series of experimental studies. This link might reflect the evolutionary connection between self-recognition and reproduction. Pinealectomy or other experimental methods which inhibit melatonin synthesis and secretion induce a state of immunodepression which is counteracted by melatonin. In general, melatonin seems to have an immunoenhancing effect that is particularly apparent in immunodepressive states. The negative effect of acute stress or immunosuppressive pharmacological treatments on various immune parameters are counteracted by melatonin. It seems important to note that one of the main targets of melatonin is the thymus, i.e., the central organ of the immune system. The clinical use of melatonin as an immunotherapeutic agent seems promising in primary and secondary immunodeficiencies as well as in cancer immunotherapy. The immunoenhancing action of melatonin seems to be mediated by T-helper cell-derived opioid peptides as well as by lymphokines and, perhaps, by pituitary hormones. Melatonin-induced-immuno-opioids (MHO) and lymphokines imply the presence of specific binding sites or melatonin receptors on cells of the immune system. On the other hand, lymphokines such as -γ-interferon and interleukin-2 as well as thymic hormones can modulate the synthesis of melatonin in the pineal gland. The pineal gland might thus be viewed as the crux of a sophisticated immunoneuroendocrine network which functions as an unconscious, diffuse sensory organ.     

Changes in connexin43, the gap junction protein of astrocytes, during development of the rat pineal gland

Abstract: The abundance of gap junctions between rat pineal astrocytes formed by connexin43 (Cx43) was studied during development. Levels and distribution of Cx43 were measured by immunoblotting and indirect immunofluorescence, respectively. The amount of Cx43 in cells located within the gland was low until about the 7th postnatal day and increased to adult values between the 14th and 21st days postpartum. Although astrocytes, recognized by their vimentin immunoreactivity, were scarce before birth, they were abundant by the 7th postnatal day suggesting that the low levels of Cx43 found at this age corresponded to a low expression of this protein. Localization of the immunoreactivity to Cx43 and vimentin showed a close correlation, indicating that mature or immature pineal astrocytes form gap junctions made of Cx43. Since Cx43 levels attained their adult values at about the time the innervation and the functional state of the gland reached maturity (2–3 weeks after birth), it is proposed that astrocyte gap junctions are involved in the function of the adult rat pineal gland.     

Response of rat pineal melatonin to calcium, magnesium, and lithium is circadian stage dependent

Abstract: Herein we documented the response of pineal melatonin production to electrolytes known to be effective on pineal function in view of a possible circadian stage dependence. We studied the release of melatonin by perifused rat pineal glands at 2 different circadian stages corresponding to the middle of the light and dark periods, i.e., respectively, 7 and 19 HALO (Hours After Light Onset, L:D = 12:12). The initial efflux rates were, as expected, much higher in the perifusates of glands removed from rats sacrificed during the dark phase than of those removed during the light phase. After 3 hr of perifusion, melatonin release reached similar levels which were found constant up to the 8th hr of perifusion, whatever the circadian stage. Perifusion of the glands with physiological concentrations for the rat of calcium (5.2 mmol/1) and magnesium (1.34 mmol/1) resulted in a stimulatory effect on the pineal glands removed from rats sacrificed in the middle of the dark period (19 HALO), whereas no effects were observed on the pineal glands removed from rats sacrificed during the light (7 HALO). Lithium (0.28 and 0.55 mmol/1) was ineffective on melatonin release in pineal glands removed 7 and 19 HALO. Our results show differences in the initial efflux rates of melatonin and in the response of perifused pineal glands to calcium and magnesium according to the circadian stage.     

Conservative management of perforated duodenal diverticulum： A case report and review of the literature

Duodenal diverticula are a relatively common condition. They are asymptomatic, unless they become complicated, with perforation being the rarest but most severe complication. Surgical treatment is the most frequently performed approach. We report the case of a patient with a perforated duodenal diverticulum, which was diagnosed early and treated conservatively with antibiotics and percutaneous drainage of secondary retroperitoneal abscesses. We suggest this method could be an acceptable option for the management of similar cases, provided that the patient is in good general condition and without septic signs.     

Presence of immunoreactive growth hormone and prolactin in the ovine pineal gland

Abstract: The use of antisera raised against bovine growth hormone (GH) and ovine prolactin (PRL) enabled the detection of related immunoreactive (ir) sequences of proteins in ovine pineal tissue. The isolation of PRL-like ir-material was accomplished using a 0.25 M ammonium sulphate (pH 5.5) extraction followed by ethanol precipitation, whereas the resulting 2.0 M ammonium sulphate (pH 7.0) precipitate contained a GH-like immunoreactivity. Gel chromatography of the GH-like immunoreactivity (Sephadex G-100) indicated the presence of several GH-like fragments ranging in the M_r range of 7,000 to 55,000. Analyses of the PRL-like ir-material found in pineal tissue on HPLC using a TSK 545-DEAE column led to the resolution into a single peak of immunoreactivity. A single peak of activity was also observed following chromatofocusing and hydrophobic interaction chromatography of the ir-peak from the TSK 545-DEAE column. The PRL-like ir-material inhibited the binding of [¹²⁵I]ovine PRL-S14 to anti-ovine PRL antibodies without showing an affinity for binding to anti-rat PRL or anti-bovine GH antibodies. Scatchard analysis of the binding of pineal PRL-like ir-material and pituitary ovine PRL-S14 to liver membranes from day-20 pregnant rats revealed similar affinity constants (K_a of 4.7 ± 0.2 × 10⁹ M^-1). In addition, the replication of Nb 2 Node rat lymphoma cells was stimulated by pineal PRL-like ir-material, an effect known to be specific for lactogenic hormones. The pineal PRL-like immunoreactivity appeared on sodium dodecyl sulfate polyacrylamide gels as a single major band of M_r 24,000. The functional status of PRL-and GH-like ir-material in the ovine pineal remains to be determined, but evidence is presented that the overall protein synthesis rate of the rat pineal responded to circulating concentrations of PRL.     

Microbiology of human immunodeficiency virus anorectal disease

PURPOSE: Individuals who are seropositive for the human immunodeficiency virus are at high risk for opportunistic infection and anorectal disorders. Little prospective information is available regarding anorectal pathogens in these patients. METHODS: One hundred sixty-three HIV-seropositive patients presented to the colorectal clinic between 1989 and 1992. Forty-seven (29 percent) patients were thought to have an infectious process and were prospectively studied using a standardized multiculture protocol. RESULTS: Mean age was 33 (range, 19–59) years. All were male; high-risk behavior accounted for 87 percent of HIV transmissions. Presenting complaints included anorectal pain (79 percent), pus per anum (28 percent), and blood per anum (26 percent). Examination revealed perianal tenderness (60 percent), condyloma (38 percent), perianal ulcers (38 percent), and anal fissures (34 percent). Sixty-six sets of cultures were performed; 28 patients had one set, 15 had two sets, and 4 had three sets. Thirty-two of these 47 patients (68 percent) had positive cultures including herpes (50 percent), cytomegalovirus (25 percent),Neisseria gonorrhoeae (16 percent), chlamydia (16 percent), acidfast bacilli (2 percent), and others (9 percent). Six of 32 patients with positive cultures had more than one organism cultured. Sixteen (50 percent) patients with positive cultures were treated medically, 8 (25 percent) were treated surgically and 8 (25 percent) were treated with both modalities. Sixty-one procedures were performed on 17 patients for condylomata. Eighteen patients had 20 procedures for abscesses, 50 percent of whom had positive cultures for other than common bowel flora; all improved. Fourteen patients underwent 33 procedures for perianal fistulas.Mycobacterium fortuitum was cultured from one patient who required 13 procedures for abscesses and fistulas. Forty-five (96 percent) patients were followed for an average of 12.5 months ±2.9 SEM (range, 1–94 months). Symptoms were improved or resolved in 22 of 32 (69 percent) patients with positive cultures and in 11 of 13 (84 percent) with negative cultures. CONCLUSIONS: Specific pathogens may often be identified in human immunodeficiency virus-seropositive patients with anorectal disorders if aggressively sought. Although patients without specific pathogens identified may be expected to improve with planned empiric treatment, positive identification allows more directed therapy.