缺血再灌注损伤后大鼠脑神经细胞巢蛋白的表达

将30只成年健康雄性SD大鼠随机分为实验组和对照组.实验组以线栓法建立大脑中动脉缺血后再灌注模型,应用免疫组化SABC法观察再灌注后各观察部位不同时间神经细胞巢蛋白(Nestin)的表达情况.发现缺血再灌注6 h,Nestin阳性细胞少量表达,1 d时数量增多,3 d时明显增多,7 d时Nestin阳性细胞数的量和形态变化最为显著.认为Nestin阳性细胞在正常成年脑组织中广泛表达,再灌注损伤后大脑各部位Nestin阳性细胞的表达呈一致性增强,各部位Nestin阳性细胞数目的 增加量在不同时间又有所不同.     

大鼠脑梗死后内源性神经干细胞的增殖及迁移

目的研究成年大鼠脑梗死后内源性神经干细胞增殖及迁移。 方法制作大鼠脑梗死模型,将其分成梗死后1、3、7、14、28 d组,对照组为假手术组。免疫组织化学方法动态检测大鼠脑内5-溴脱氧尿苷嘧啶(BrdU)、巢蛋白(Nestin)的表达。 结果与正常对照组相比,室下区及海马BrdU和Nestin阳性细胞在脑梗死后1 d开始增加(P<0.05),7d达到高峰,14 d后开始下降,但仍高于正常水平(P<0.05),28 d后接近正常水平;室下区及海马BrdU和Nestin阳性细胞经胼胝体向对侧迁移。 结论脑梗死可激活内源性神经干细胞原位增殖及迁移。     

清热化瘀方对脑缺血再灌注损伤大鼠神经干细胞增殖的影响

目的 观察清热化瘀方对大鼠脑缺血再灌注损伤后神经干细胞增殖的影响.方法 90只大鼠随机分为假手术组、模型组、清热化瘀组3组,采用线栓法制作局灶性脑缺血大鼠(MCAO)模型,观察大鼠缺血损伤后缺血半暗带区的细胞增殖.采用免疫组化法观察脑缺血再灌注后2 d、4 d、7 d、14 d、21 d五个时间点巢蛋白(Nestin)及5-溴脱氧尿苷(BrdU)免疫活性的变化.结果 清热化瘀组大鼠在第2天、第4天、第7天的神经功能缺损评分明显优于模型组(P<0.05);模型组缺血半暗带Nestin和BrdU阳性表达分别于再灌注7 d和14 d达高峰(P<0.05),以后表达逐渐减弱;清热化瘀组可以明显增加缺血再灌注后第4天、第7天、第14天大鼠的Nestin阳性细胞表达(P<0.05);并可以明显增加缺血再灌注后第7天、第14天、第21天大鼠缺血半暗带的BrdU阳性细胞表达(P<0.05).结论 脑缺血损伤可激活神经干细胞,促使其增殖和表达.清热化瘀方可以促进MCAO大鼠缺血半暗带神经干细胞的增殖.     

腹腔注射尿酸对脑缺血缺氧损伤保护作用的实验研究

目的 探讨尿酸对脑缺血缺氧损伤的保护作用及机制.方法 将7日龄新生SD大鼠随机分为对照组、模型组及尿酸组（UA）各15只,模型组及UA组均参照Rice-Vannucci方法制备脑缺血缺氧模型,其中UA组模型制作完毕后6 h腹腔注射UA 62.5 mg/kg;对照组常规母鼠喂养.术后24 h三组均过量麻醉、快速断头取脑,观察脑组织含水量,并经HE染色观察脑皮层和海马损伤情况.结果 UA组脑组织含水量显著少于模型组,脑皮层和海马损伤程度明显轻于模型组.结论 新生大鼠脑缺血缺氧损伤后6 h腹腔注射UA具有神经保护作用,可能机制为抑制氧化损伤;此为临床治疗新生儿缺血缺氧性脑病提供了理论基础.     

大鼠颅脑损伤后巢蛋白变化的实验研究

目的观察大鼠颅脑损伤后不同时期损伤周边区皮质神经细胞凋亡现象，为颅脑外伤的临床治疗提供理论依据。方法将SD大鼠分为正常对照组（5只）和损伤组（90只），损伤组用侧位液压冲击法制成颅脑损伤动物模型，进一步分为轻、中、重三组，采用免疫组织化学技术检测各组损伤后3、6、24．72h．1、2w大鼠伤灶周边皮层的巢蛋白表达情况。结果损伤3h后伤灶边缘皮层开始出现巢蛋白阳性细胞，随后逐渐增加，1w达到高峰，随着时间的推移其表达逐渐下降，假手术组各时间点伤处边缘皮层也可见到少量巢蛋白阳性细胞，但低于损伤组，具有统计学意义（P〈0．05）。结论颅脑外伤可导致损伤周边区大脑皮层阳性巢蛋白神经细胞的增多，损伤后1w为巢蛋白阳性细胞表达的高峰时间。     

局灶性脑缺血后神经干细胞增殖及GDNF表达变化的实验研究

目的研究局灶性脑缺血后大鼠脑内神经干细胞(NSCs)增殖情况及与胶质细胞源性神经营养因子(GDNF)表达变化的关系。方法将24只SD大鼠随机分为假手术组和模型1组、模型2组、模型3组各6只,后三组用大脑中动脉栓塞法(MCAO)制备局灶性脑缺血模型,分别于术后3、7、14 d处死;假手术组除不以尼龙线阻塞大脑中动脉起始部外,其他操作与模型1组相同。采用免疫组化染色法观察5-溴脱氧尿嘧啶核苷(BrdU)标记的细胞及巢蛋白(Nestin)、GDNF阳性细胞表达,BrdU/Nestin免疫荧光双标法观察NSCs变化。结果模型2组损伤侧脑室下层(SVZ)BrdU阳性细胞显著多于假手术组(P<0.05),模型3组与假手术组相似;模型2、3组皮层梗死灶周围BrdU阳性细胞均显著多于假手术组(P<0.01、0.05);模型1、2组皮层梗死灶周围Nestin阳性细胞均显著多于假手术组(P均<0.01),BrdU/Nestin免疫荧光双标显示BrdU阳性细胞几乎均为Nestin阳性;模型2、3组皮层梗死灶周围GDNF阳性细胞均显著多于假手术组(P均<0.05)。结论大鼠局灶性脑缺血后3～14 d内源性NSCs增殖加快,GDNF表达增加可能对其起促进作用。     

神经干细胞移植治疗脑出血大鼠模型时间窗的研究

目的研究神经干细胞移植治疗脑出血的时间窗。方法分离、培养大鼠海马神经干细胞，经Nestin免疫荧光鉴定，Brdu标记；25只SD大鼠制作脑出血模型，随机分为5组（1、2、3、4d移植组、模型组），分别在造模后第1、2、3、4d移植Brdu标记的神经干细胞于脑出血大鼠的患侧侧腔室下区（SVZ），模型组不移植，测试大鼠运动功能，14d时处死，取全脑固定、冰冻切片、Brdu免疫荧光染色，Brdu阳性细胞计数。结果各移植组大鼠神经功能缺损明显改善，2d移植组明显优于其他组（P〈0．01）；2d移植组Brdu阳性细胞数明显高于其它组（P〈0．05）。结论神经干细胞移植治疗脑出血能有效改善脑出血动物运动功能；脑出血后2d可能是神经干细胞移植的最佳时间点。     

胎鼠神经干细胞移植对阿尔茨海默病大鼠海马区的GFAP与S-100β表达的影响

目的探讨胎鼠神经干细胞移植对阿尔茨海默病(AD)大鼠海马区的GFAP与S-100β表达的影响。方法通过切断Wistar大鼠海马穹窿伞制备AD模型大鼠。我们将大鼠随机分为神经干细胞移植组、生理盐水对照组和正常组。培养胎鼠的神经干细胞,将其移植入AD模型大鼠脑内,并用等量生理盐水脑内注射对比观察。1个月后处死大鼠,进行免疫组织化学实验检测鼠海马区的GFAP与S-100β的表达。结果神经干细胞显示巢蛋白阳性。各组实验鼠海马区有GFAP的阳性表达,生理盐水对照组与其他各组之间有显著差异(P<0.01)。对各组大鼠在海马区均可见棕褐色的S-100β阳性细胞存在,部分细胞呈细胞质性染色阳性。生理盐水对照组AD模型大鼠的海马区切片上可见大量阳性细胞染色,与其他各组比较差异有显著性(P<0.01)。结论神经干细胞移植可以促进AD模型大鼠脑内神经元更好的恢复。     

大鼠海马神经干细胞的原代培养方法

目的探索总结一种简单、高效的大鼠海马神经干细胞原代培养方法。方法选取新生24 h的SD大鼠,剥离出双侧海马,用机械吹打法制成细胞悬液,加入培养液,接种于培养皿中,3 d后半量换液,5~7 d后可传代。每天在荧光倒置显微镜下观察细胞的生长状况,采用免疫荧光法对第2代神经干细胞进行鉴定。结果本方法能准确分离出海马组织,培养出的神经干细胞中无其他混杂的细胞生长,经鉴定呈兔抗巢蛋白(Nestin)阳性和5-嗅-2-脱氧尿苷(BrdU)阳性。结论运用本方法获得的海马神经干细胞纯度高、细胞活力高、增殖能力强。     

神经节苷脂联合参附注射液对新生大鼠缺氧缺血性脑损伤的保护作用

目的探讨神经节苷脂(GM1)联合参附注射液对新生大鼠缺氧缺血性脑损伤(HIBD)的神经保护作用。方法将新生7d SD乳鼠90只随机分为假手术组、缺血缺氧组(HIBD组)、缺血缺氧+GM1干预组(GM1组)、缺血缺氧+参附干预组(参附组)及缺血缺氧+GM1及参附干预组(合用组)。在给药后不同时间点,采用免疫组化法半定量测定脑组织中Bcl-2及Bax含量。结果假手术组海马CA1区可见少量Bcl-2及Bax表达;HIBD后该区Bcl-2及Bax表达均增加,且Bax表达较明显;分别应用GM1、参附注射液治疗后该区Bcl-2表达进一步增加,而Bax蛋白水平下降;联合应用两种药物治疗后该区Bcl-2表达增加、Bax表达下降的趋势更为明显。结论新生大鼠脑缺氧缺血后腹腔注射神经节苷脂或参附注射液均可不同程度减轻大鼠脑损伤,两种药物联合应用对其脑损伤的改善作用更为明显。     

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.     

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.     

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.     

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.