自然杀伤T细胞在急性肾损伤小鼠肾纤维化中的作用

目的:评价自然杀伤T细胞在急性肾损伤小鼠肾纤维化中的作用。方法:清洁级健康雄性C57BL/6小鼠24只,8～10周龄,体重20～30 g,采用随机数字表法分为4组( n＝6)：对照组(C组)、急性肾损伤组(A组)、对照组+CD1d抗体组(C-MA组)和急性肾损伤+CD1d抗体组(A-MA组)。腹腔注射叶酸2...     

自然杀伤T细胞失能的相关研究进展

自然杀伤T细胞同时表达自然杀伤细胞和T细胞受体,发挥免疫调节和细胞毒作用.相关研究证实激活自然杀伤T细胞可以抑制肿瘤进展,但短暂活化的自然杀伤T细胞常常因为未知原因迅速进入失活状态.本文就目前自然杀伤T细胞失能可能机制及应对措施的研究进展作一综述.     

肾结核患者尿液抗酸杆菌与自然杀伤T细胞的关系

目的探索肾结核患者尿液抗酸杆菌与自然杀伤细胞T（naturalkillerTcells,NKT）功能的相关性。方法将54例肾结核患者分为活动期组（n=28）及非活动期组（n=26）,并选取20名健康者作为对照;采用多色流式细胞仪检测外周血NKT细胞百分含量,同法检测NKT胞内细胞因子自介素-4（IL-4）、干扰素-γ（IFN-γ）及白介素-12（IL-12）含量;采用荧光定量PCR法检测尿液抗酸杆菌TB-DNA含量,使用ELISA法检测尿液细胞因子IL-4、IFN-γ及IL-12含量。结杲在外周血NKT及其细胞因子、以及尿液细胞因子含量比较方面,均为活动期患者〈非活动期患者〈健康人;而尿液TBDNA含量比较显示活动期患者〉非活动期患者〉健康人,以上任意两组间均有显著性差异（P〈0．05）。相关性分析表明外周血NKT与尿液TB-DNA含量呈显著负相关（r=-0．878,P〈0．0001）。结论肾结核患者NKT细胞功能明显降低,且与是否处于活动期密切相关;反映病灶局部的尿液抗酸杆菌的含量与反映全身免疫功能的NKT细胞数量级功能均有紧密联系。     

手术前后前列腺癌患者自然杀伤T细胞的变化及意义

【摘要】〓目的〓通过自然杀伤T细胞(NKT)评价经腹膜外腹腔镜前列腺癌根治术(ELRP)的免疫调节效果。方法〓收集35例前列腺癌患者,以35例名健康对照组作为对照,采用流式细胞术检测外周血NKT及其细胞因子γ干扰素(IFN-γ)和白介素4(IL-4)含量。前列腺癌患者接受ELRP术后2周再次检测上述指标观察其变化。结果〓前列腺癌患者NKT百分含量、IFN-γ及IL-4的平均荧光强度分别为5.92±1.47、4.23±1.90及2.15±1.04,明显低于对照组的12.53±4.16、8.94±2.76及5.86±1.97(P<0.05);前列腺癌患者术后三者分别回升为9.13±2.69、6.19±2.03 及3.37±1.28,与术前差异有统计学意义(P<0.05),且均以IFN-γ的平均荧光强度改变较IL-4明显。结论〓NKT及其细胞因子数量下降是前列腺癌患者的一个重要免疫学特征,ELRP术可改善前列腺癌患者的细胞免疫功能紊乱。     

不育男性精液中精子及生精细胞的形态学检查

不育男性精液中精子及生精细胞的形态学检查张卫青，孙明祥，张同金，汪涛，孙天玉关键词不育男性；精液；精子精液中精子及生精细胞的形态学检查，是评价男性生育能力不可缺少的部分。形态检查染色方法很多，但都比较繁杂，不适于短时间内做大量标本。我们把甲苯胺蓝、副...     

精索静脉曲张患者精液质量及其不育患者手术前后精液变化的观察

目的观察精索静脉曲张（VC）患者的精液质量和精子形态学改变，以及VC不育患者手术前后精液的变化。方法121例VC患者精液按WHO标准常规分析并对精子形态学进行评价，23例健康男性精液检查结果作为对照。并对21例VC不育患者术前及术后的精液进行检测分析。结果121例VC患者的精子密度、（A＋B）级活动力精子（％）、活率、有效精子数、活动精子数、活力指数以及正常形态精子比例较对照组明显降低（P〈0．01）；畸形精子中小头、锥形头和无定形头精子数较对照组增多（P〈0．01）。21例VC不育患者手术后精子质量和精子形态学较术前明显改善。结论VC可以引起精液质量下降导致不育，精子形态学分析是判定VC患者精子受损的一个敏感指标，手术能有效地改善精液质量。     

Clusterin蛋白在男性不育症睾丸组织中的表达及意义

目的 探讨Clusterin蛋白在男性不育症患者睾丸组织中的表达及意义.方法 采用免疫组化染色法和Western blot法检测了10例正常睾丸组织,8例梗阻性不育患者睾丸组织,13例非梗阻性不育患者睾丸组织标本中Clusterin蛋白的表达水平.结果 免疫组化染色结果表明,3组睾丸组织中Clusterin蛋白都有表达,.正常睾丸组织、梗阻性不育患者睾丸组织、非梗阻性不育患者睾丸组织中阳性或强阳性表达率分别为100%(10/10),87.5%(7/8),46%(6/13),非梗阻性不育睾丸组织中Clusterin蛋白表达水平显著低于正常睾丸组织(P<0.05).Western blot法检测结果也表明,该蛋白在非梗阻性不育睾丸组织中的表达较正常睾丸组织显著降低,而在梗阻性不育睾丸组织中的表达无显著性降低.结论 Clusterin蛋白的表达降低与非梗阻性不育症的发生密切相关.     

杭州市萧山区不育患者精液质量及动态参数分析

目的 研究杭州市萧山区男性精液质量及各项精子动态参数在评价男性生殖方面的临床应用价值.方法 对本院2 583例男科门诊就诊的患者精液用计算机辅助精液分析(CASA)系统进行常规指标和精子动态参数检测.结果 2 583例精液标本中,无精子和死精子142例,其余按照WHO正常精液评判标准,七项指标均正常的精液1 268例,占49.09%;一项指标异常314例,占12.16%;二项指标异常216例,占8.36%;≥三项以上指标异常643例,占24.89%.1173例异常精液中(不包括死精子和无精子),活力异常749例,检出率63.85%;活率异常608例,检出率51.83%;密度异常406例,检出率34.61%.活力、活率异常组与正常组动态参数比较,曲线速度(VCL)、直线速度(VSL)、直线性(LIN)、摆动性(WOB)、平均路径速度(VAP)、平均移动角度(MAD)、前向性(STR)下降明显;密度异常组中VCL、VSL、LIN、WOB、VAP与正常组比较有统计学差异;鞭打频率(BCF)在3个异常组中均呈上升趋势.结论 杭州市萧山区男性精子活率、活力和密度异常所致不育的情况较精液量、液化时间、酸碱度异常及畸精子症致不育的情况更常见;精子动态参数是评估精液质量的良好指标,为男性不育的诊断、治疗和研究提供了充分的依据.     

干扰素γ诱导人肝癌细胞表达HLA-E逃避NK细胞杀伤

目的探讨干扰素γ（IFN-γ）诱导人肝癌细胞表达非经典人类白细胞抗原HLA-E,及对NK细胞肿瘤杀伤的影响。方法用不同终浓度的IFN-γ分别体外持续诱导人肝癌细胞系（BEL-7402）72h。用细胞免疫组织化学、Western-blot、逆转录PCR和流式细胞技术,检测各组细胞HLA-E基因产物的表达及各组的NK细胞杀伤率。结果与空白对照相比,IFN-γ浓度100～1000U／ml）诱导BEL-7402细胞表达HLA-E基因产物（抗原、mRNA和蛋白）明显增多（P〈0．01）,细胞表面HLA-E抗原相应增多（P〈0．01）。细胞表达HLA-E基因的强弱与IFN-γ诱导浓度有关。高浓度IFN—γ（500-1000U／ml）诱导细胞HLA-E阳性率、表达强度高于低浓度诱导（100-400U／ml）。最适宜的IFN-γ浓度为500U／ml,高浓度组的NK细胞杀伤率更低（P〈0．01）。结论高浓度IFN-γ（500～1000U／ml）诱导人肝癌细胞HLA-E基因高表达,通过非经典HLA-Ⅰ（HLA-E）途径逃避NK细胞免疫监视。     

High percentage of apoptotic spermatozoa in ejaculates from men with chronic genital tract inflammation

Silent chronic inflammation of genital tract (CIGT) is considered as a major contributing factor to male fertility disorders. Within CIGT, inflammatory cytokines such as TNF-alpha might induce spermatozoal apoptosis, which in turn has been shown to have a negative influence on the sperm-oocyte penetration capacity. Thus, the aim of this study was to investigate spermatozoal apoptosis in patients with fertility disorders and signs of CIGT. Apoptosis of spermatozoa was determined by expression of annexin V using flow cytometry. Apoptotic spermatozoa were further discriminated from necrotic spermatozoa by 7-amino-actinomycin (7AAD). Ejaculates of patients showing signs of CIGT (P-CIGT) such as high polymorphonuclear (PMN) elastase were compared to control ejaculates of patients lacking signs of CIGT (CON). Thereby we detected a significant correlation between PMN elastase and TNF-alpha in the seminal plasma and apoptotic spermatozoa. Furthermore, we demonstrated a significantly higher percentage of apoptotic spermatozoa in the ejaculate of P-CIGT compared to CON. Interestingly, a significantly higher percentage of apoptotic spermatozoa was detected in the swim-up fraction demonstrating motility of apoptotic spermatozoa. This is in line with the missing correlation of apoptotic spermatozoa and motility. In conclusion, patients with signs of CIGT display high numbers of apoptotic spermatozoa with progressive motility, which might have an impact on reproductive techniques such as intrauterine insemination or in-vitro fertilisation.     

Reduction of natural killer and natural killer T cells is not protective in cisplatin-induced acute renal failure in mice

Aims: A recent report showed that fractalkine (CX3CL1), which functions as both a potent chemoattractant and adhesion molecule for monocytes and natural killer (NK) cells was significantly increased in cisplatin‐induced acute renal failure (CisARF) in mice. Therefore, we developed the hypothesis that increased CX3CL1 expression in CisARF initiates NK cell infiltration in the kidney. The aim of the present study was to determine the role of NK cells in CisARF in mice. Methods: Time course of pan‐NK positive cells in CisARF was investigated by using immunohistochemistry (IHC) for CD49b. Pan‐NK positive cells were reduced by using anti‐NK1.1 mAb. The model of pan‐NK positive cells reduction was confirmed by flow cytometry of the spleen and IHC of the kidney. The expression of granzyme A and caspase‐1 was examined, and the activity of caspase‐1 was also determined. We performed a study on whether there was significant protection of renal function after reduction of pan‐NK positive cells. Results: (i) Infiltration of pan‐NK positive cells was prominent on day 3 after cisplatin administration. (ii) granzyme A expression was significantly increased in CisARF and CisARF+NK1.1 Ab compared to vehicle. (iii) Caspase‐1 expression and activity was significantly increased in CisARF mice compared to vehicle and CisARF+NK1.1 Ab. (iv) Reduction of pan‐NK positive cells was not protective in cisplatin‐induced acute renal failure in mice. Conclusions: Although infiltration of pan‐NK cells was significantly increased in CisARF, reduction of infiltration of pan‐NK cells into the kidney was not protective against CisARF in mice.     

Seminal macrophages in ejaculates from men with couple infertility

Seminal macrophages are occasionally reported though their relevance in the evaluation of human ejaculate is unknown. Activated macrophages, engaging in sperm phagocytosis (spermiophages), might represent a marker of innate immunosystem activation. We investigated whether the presence of spermiophages in non-leukocytospermic ejaculates from men complaining for couple infertility is associated with altered sperm features. Four hundred and thirty-four ejaculates were retrospectively analysed after excluding samples with antisperm antibodies, or a leukocyte number ≥1 × 10⁶/mL. Semen quality was compared in samples with or without spermiophages detected with transmission electron microscope. Presence of spermiophages, observed in 27% of ejaculates, was associated with a decreased number of sperm total count ( p  < 0.0001), of sperm forward motility ( p  = 0.048), and to an increased fraction of degenerating sperm ( p  = 0.0002) compared to ejaculates without spermiophages. A low number of total ejaculated sperm and an increased number of degenerating sperm independently predicted the presence of spermiophages (odds ratio 1.72; 95% confidence intervals 1.10 to 2.28 and odds ratio 1.85; 95% confidence intervals 1.19 to 2.88 respectively). Data demonstrate that activated macrophages, a marker of the innate immunosystem activation, are frequently observed in non-leukocytospermic ejaculates of men suffering for couple infertility and this may be associated with altered sperm parameters. Ultrastructural analysis gives qualitative informations, hence sensitive quantitative tests should better define the association between semen activated macrophages and oligoasthenozoospermia and the possible relevance of this finding in the clinical evaluation of the male partner of couple infertility.     

Expansion of natural killer cells in patients with head and neck cancer: detection of "noninhibitory" (activating) killer Ig-like receptors on circulating natural killer cells

BACKGROUND: In a group of patients with head and neck cancers (H&NC), the expansion of the population of CD3-,CD16+ natural killer (NK) cells in the peripheral blood was studied. METHODS: Cytofluorimetric analysis of the expression of killer Ig-like receptors (KIR, namely p58.1, p58.2, p58.3, p70, and p140) and CD94-NKG2a was performed. Cytolytic activities were studied using 51Cr release assay. T and NK cell cloning was performed using limiting dilution culture conditions. Cytokine production was analyzed using commercial enzyme immunoassays. RESULTS: Phenotypic analysis showed that the expanded populations were heterogeneous. Even in the presence of a large number of circulating NK cells, "nonspecific" cytolytic capacities were heavily reduced, whereas cytolytic capacity related to T cells was virtually normal. Unlike NK cell clones derived from healthy donors, most NK cells derived from H&NC patients expressed surface "activating" NK cell receptors (KAR) for HLA, detected by use of a redirected cytolytic assay. Analysis of the CD4+ subpopulation at the clonal level demonstrated that they had a severe proliferative defect. CONCLUSION: These experimental data indicated that H&NC patients have a polyclonal expansion of functionally deficient NK cells expressing KAR. In addition, the proliferative capacity of patients' "helper" cells was strongly inhibited, thus accounting for a severe impairment of cytolytic activity of the expanded NK cells.     

Elastase as an indicator of silent genital tract infection in infertile men

Due to the absence of clinical symptoms, silent genital tract inflammation can be diagnosed only by laboratory tests. In this study we have evaluated seminal plasma elastase levels, using an immunoabsorbent assay, in a group of 84 infertile men. Seminal plasma levels of elastase were correlated with the number of white blood cells in the ejaculate, the number of peroxidase-positive leucocytes and with sperm culture. A high number of leucocytes (greater than 10) and a significantly higher number of men with peroxidase-stained leucocytes exceeding 10(6)/ml was found in a group of men with elastase levels greater than 250 ng/ml. There was a significant correlation between sperm culture results and elastase levels, most men with negative sperm culture having a lower seminal plasma elastase level. Following the treatment with antibiotics of men with an elevated elastase level, sperm parameters improved in 67% of those in whom elastase levels were lowered after treatment. In those men with persisting elevated levels of elastase improvement of sperm parameters was found in only 10%. It is concluded that an elevated level of elastase is a sensitive indicator of asymptomatic genital tract infection and that a single determination gives a reliable criterion and relatively exact quantification of infection.     

The role of natural killer T cells in costimulation blockade‐based mixed chimerism

Distinct lymphocyte populations have been identified that either promote or impede the establishment of chimerism and tolerance through allogeneic bone marrow transplantation (BMT). Natural killer T (NKT) cells have pleiotropic regulatory properties capable of either augmenting or downmodulating various immune responses. We investigated in this study whether NKT cells affect outcome in mixed chimerism models employing fully mismatched nonmyeloablative BMT with costimulation blockade (CB). The absence of NKT cells had no detectable effect on chimerism or skin graft tolerance after conditioning with 3Gy total body irradiation (TBI), and a limited positive effect with 1Gy TBI. Stimulation of NKT cells with alpha‐galactosylceramide (alpha‐gal) at the time of BMT prevented chimerism and tolerance. Activation of recipient (as opposed to donor) NKT cells was necessary and sufficient for the alpha‐gal effect. The detrimental effect of NKT activation was also observed in the absence of T cells after conditioning with in vivo T‐cell depletion (TCD). NKT cells triggered rejection of BM via NK cells as chimerism and tolerance were not abrogated when NKT cells were stimulated in the absence of both NK cells and T cells. Thus, activation of NKT cells at the time of BMT overcomes the effects of CB, inhibiting the establishment of chimerism and tolerance.     

The relationship of natural killer cells to metastasis of a transplantable prostate adenocarcinoma

Natural killer (NK) cells have been proposed to play a significant role in the inhibition of metastasis. The prostate adenocarcinoma (PA-11) in the Lobund-Wistar (L-W) rat provides a unique model of spontaneous metastasis in which to study NK response. Cultured PA-11 tumor cells were shown to be resistant to NK lysis in vitro, and enhancement or inhibition of NK reactivity in vivo using drugs or antiserum did not change the rate or extent of metastasis evident at autopsy. Exposure to PA-11 tumor cells, supernatants from cultured tumor cells, or sera from rats with advanced PA-11 in vitro did not result in inhibition of NK activity. Exposure to PA-11 tumor cells in vivo also did not cause suppression of NK activity. These data indicate that, in the PA-11/L-W system, metastasis is independent of NK activity.     

Low-dose remifentanil infusion does not impair natural killer cell function in healthy volunteers

Background. Mu opioid agonists suppress natural killer (NK)cell activity in animal models. Studies in human volunteers,however, have yielded conflicting results, with morphine suppressingand fentanyl increasing NK cell activity. This study evaluatedthe effect of a constant 8-h infusion of remifentanil on NKcell number and function in human volunteers. Methods. After IRB approval and informed consent was obtained,10 healthy volunteers underwent an 11 pm to 7 am infusion ofsaline, and at least 1 week later an infusion of 0.02–0.04µg kg^–1 min^–1 remifentanil. Blood was collectedat 7 am for measurement of NK cell cytotoxicity using a ⁵¹Crrelease assay and measurement of NK cell number using fluorescentflow cytometry. Results. Median and range of the total NK cell cytoxicity (KUml^–1) was 745.0 (498.3–1483.6) on the control morningand 818.6 (238.5–1454.5) on the morning following theremifentanil infusion. Neither the number of NK cells ml^–1(2.5x10⁵ (1.4x10⁵–4.2x10⁵) vs 2.7x10⁵ (1.1x10⁵–4.4x10⁵))nor the cytotoxicity per 1000 NK cells (KU 1000 NK cells^–1)(3.0 (1.8–5.2) vs 2.9 (0.9–6.7)) changed betweenthe control and remifentanil conditions. Conclusions. An 8-h infusion of remifentanil did not affectNK cell activity in normal volunteers. This result differs fromprevious findings of morphine-induced NK cell activity suppressionand fentanyl-induced NK cell activity enhancement in normalvolunteers. Br J Anaesth 2003; 91: 805–9