MRL/lpr鼠骨髓间充质干细胞对T淋巴细胞作用的研究

目的 探讨狼疮鼠（MRL／lpr）及正常鼠（C57BL／6）骨髓间充质干细胞（MSCs）体外生物学特性、对T淋巴细胞作用的影响。方法 采用直接贴壁筛选法分离培养扩增两种鼠系骨髓MSCs，流式细胞术鉴定MSCs表面标记物，观察MSCs生长状况并绘制生长曲线；取C57BL／6鼠脾脏细胞，经尼龙毛柱分选CD3T淋巴细胞．经刀豆蛋白A（ConA）刺激，取正常及狼疮鼠MSCs或MSCs培养上清与T细胞共培养72h，然后羧基荧光素二醋酸盐琥珀酰亚胺酯（CFSE）染色测定T细胞增殖，流式细胞术检测共培养前后T细胞活化及T细胞的凋亡率。结果 第2代（P2）后狼疮鼠MSCs生长增殖快于正常鼠MSCs；正常及狼疮鼠MSCs共培养均可抑制正常鼠T细胞的增殖（P〈0．05），与MSCs上清共培养对正常鼠T细胞增殖无抑制作用（P〉0．05）；与两种MSCs或上清共培养均可降低正常T细胞的活化（P〈0．05）及凋亡率（P〈0．05）。对正常T细胞增殖、活化、凋亡的影响，狼疮鼠MSCs与正常鼠MSCs间差异无统计学意义（P〉0．05）。结论 狼疮鼠MSCs在体外生长上存在异常．但对T细胞增殖、活化、凋亡免疫调节无异常。     

人骨髓间充质干细胞治疗再生障碍性贫血的机制研究

目的探讨在体外骨髓间充质干细胞(MSC)治疗再生障碍性贫血(AA)的机制。方法2003-02~2005-04安徽医科大学第一附属医院血液内科从人骨髓分离培养MSC,用尼龙棉柱分离T淋巴细胞。分别以不同数量MSC加入到植物血凝素(PHA)刺激的AA患者T淋巴细胞增殖体系中,用MTT还原法测定MSC对AA患者T细胞增殖的影响,通过流式细胞术计算CD3+T细胞CD25(IL-2R)的表达率。结果当MSC为2×104/孔(A组)和1×104/孔(B组)时,与单独培养的AA患者T淋巴细胞(对照组)相比,MSC对AA患者T淋巴细胞增殖及CD25的表达呈明显抑制作用,差异有统计学意义(P<0·01)。当MSC为1×103/孔(C组)时,无统计学意义。结论MSC抑制AA患者T细胞的活化与增殖,且这种抑制作用具有数量依赖性。     

环孢霉素A治疗再生障碍性贫血的现状与前景

尽管再生障碍性贫血（AA）病理机制呈现高度异质性，但近年来大量的实验室及临床研究结果表明，免疫介导的造血抑制是最常见的病理机制[1，5]。作为特异性免疫抑制剂，环孢霉素A（CsA）无论单用或与其它免疫抑制剂联合应用，其治疗AA疗效确切性已获公认。本文就CsA治疗AA的研究现状作一综述。1AA免疫病理机制近年来，有关AA免疫病理机制的研究已获突破性进展。研究结果证实，AA骨髓造血功能衰竭主要与活化的细胞毒性T细胞（CTL）有关，CTL的活化与增殖主要局限于骨髓这一靶器官[2]。AA活化的CTL体外表现出明显造血抑制活性，对…     

香丹注射液体外诱导大鼠MSCs分化为多巴胺能神经元效果观察

目的探讨香丹注射液体外诱导大鼠骨髓间充质干细胞（MSCs）定向分化为多巴胺（DA）能神经元的效果。方法体外分离、扩增和鉴定大鼠骨髓MSCs。碱性成纤维细胞生长因子（bFGF）预诱导24h后香丹注射液诱导3h，采用免疫细胞化学法检测神经元特异性烯醇化酶（NSE）、微管相关蛋白2（MAP2）、胶质纤维酸性蛋白（GFAP）、酪氨酸羟化酶（TH）、增殖细胞核抗原（PCNA）、Bcl-2表达。结果诱导后MSCs分化为具有典型神经元形态的细胞，NSE、MAP2和TH呈高表达，PCNA表达明显减弱。结论香丹注射液诱导大鼠MSCs分化为DA能神经元效果较好。     

血管内皮生长因子对骨髓间充质干细胞分离培养的诱导分化作用

目的　结合骨髓间充质干细胞与血管内皮生长因子（VEGF）的优点,探讨VEGF对其体外诱导分化的作用。为缺血性疾病的治疗提供新思路。方法　成人新鲜骨髓,Percoll梯度分离法培养,单克隆培养法分离传代扩增骨髓基质细胞（MSCs）。在大肠杆菌DH5α中扩增和提取,纯化、克隆pcDNA3.0-VEGF165质粒。脂质体法转染MSCs。流式细胞术检测诱导前后MSCs免疫表型变化。一抗VEGF和CD31（1∶50）,FITC标记的VEGF二抗和cy3标记的CD31二抗与细胞共同孵育,甘油封片,在激光扫描共聚焦显微镜下观察细胞染色情况。结果　转染前表达CD44（75.86%）、CD31（5.63%）。转染后CD44表达明显下调（40.18%）,CD31则明显上调（56.20%）。FITC标记后VEGF抗体使细胞显现绿色荧光,cy3标记的CD31抗体使细胞显红色荧光。结论　转染VEGF后骨髓间充质干细胞向内皮细胞分化且有分泌VEGF的功能,为干细胞植入及血管再生起到指导作用。     

MSCs诱导移植物免疫耐受的研究进展

上世纪60-80年代，Friedenstein和他的同事发现骨髓细胞培养后有一种贴壁生长的成纤维样细胞可以在体内成骨，称之为骨髓间充质干细胞（MSCs）。随后发现，作为多系细胞的祖细胞，MSCs可以从脂肪组织、胎肝和脐带等多种组织中分离出来，在体内外可以分化为多种间充质组织，故可用于再生医学领域。另外，MSCs还具有免疫调节活性，在体外阻断T细胞增殖，     

骨髓间充质干细胞及基因治疗心血管疾病的研究进展

骨髓间充质干细胞（mesenchymal stem cells，MSCs）是存在于骨髓中，除造血干细胞外的另一类早期前体细胞；其来自基质，也称骨髓基质干细胞。1987年Friedenstein等在分离大鼠骨髓成骨祖细胞时，首次从骨髓基质中鉴定出一种非造血系成体多能干细胞，并实验证实在体外这类细胞能分化成各种间质细胞，包括成骨细胞、脂肪细胞、软骨细胞和肌肉细胞等，因而称之为MSCs。     

AA和MDS患者骨髓CD+34细胞表面GM-CSFR的表达及意义

目的 探讨再生障碍性贫血（AA）和骨髓增生异常综合征（MDS）患者骨髓CD34^＋细胞表面粒细胞-巨噬细胞集落刺激因子受体（GM—CSFR）的表达情况。方法 用流式细胞术（FCM）和单克隆抗体技术检测14例AA、23例MDS和8例非血液病患者骨髓CD34^＋细胞表面的GM—CSFR表达率。结果 MDS组骨髓CD34^＋细胞表面GM-CSFR的表达率明显高于对照组及AA组（P〈O．05），而AA组与对照组间差异无统计学意义（P〉0．05）。结论 AA患者造血干细胞没有质的缺陷；造血干细胞异常可能是MDS的主要发病机制之一。     

间充质干细胞的免疫调节作用及其临床应用

近年来对间充质干细胞（mesenchymal stem cells，MSCs）的研究越来越多，MSCs可以从骨髓、脂肪组织、肾及各种胎儿组织中提取，并能横向分化成成骨细胞、脂肪细胞和神经细胞等不同胚层的细胞。MSCs通过产生一系列细胞因子，作用于抗原提呈细胞、T细胞和NK细胞等免疫细胞而抑制机体免疫功能的发挥。基于上述多潜能分化和免疫抑制特性，MSCs可以用于移植、诱导移植耐受和治疗自身免疫性疾病。     

系统性红斑狼疮患者骨髓间质干细胞对B淋巴细胞免疫抑制作用的初步研究

目的：探讨系统性红斑狼疮（SLE）患者骨髓间质干细胞（MSCs）体外对B淋巴细胞的免疫调节作用是否存在异常。方法：采用直接贴壁法分离培养正常人及SLE患者MSCs。免疫磁珠法分选正常人外周血B淋巴细胞,与MSCs共培养,流式细胞术检测B细胞凋亡及表面标志表达,3H掺入法检测B细胞增殖,酶联免疫吸附法（ELISA）检测培养上清中免疫球蛋白IgG、IgM和IgA水平。结果：①B细胞＋正常MSCs共培养组B细胞增殖（4307±308）cpm较单纯B细胞培养组（7059±346）cpm降低（P〈0.05）;②B细胞＋正常MSCs共培养组B细胞凋亡率（14±7）%低于单纯B细胞培养组（25±8）%（P〈0.05）;③B细胞＋正常MSCs共培养组B细胞表面CD86表达率（53±5）%较单纯B细胞培养组（66±10）%低（P〈0.05）;④与MSCs共培养后,B细胞分泌免疫球蛋白IgG、IgM和IgA较单纯B细胞培养显著减少（P〈0.05）;⑤B细胞＋狼疮MSCs共培养组与B细胞＋正常MSCs共培养组比较,B细胞增殖、凋亡、CD86表达和Ig分泌两组间差异无统计学意义。结论：MSCs在体外对B细胞有抑制作用,SLE患者骨髓MSCs与正常MSCs相比,对B细胞的表面刺激分子表达、增殖、Ig分泌有相似的抑制作用。     

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.     

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.     

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.     

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.