Li R K, Mitchell T G
Department of Microbiology, Duke University Medical Center, Durham, NC 27710, USA.
J Med Vet Mycol. 1997 Sep-Oct;35(5):327-34.
Lewis rat alveolar macrophages (AM) were harvested and exposed in vitro to Cryptococcus neoformans to investigate the induction of inflammatory cytokines. AM in tissue culture wells were incubated with viable yeast cells of C. neoformans or the capsular polysaccharide, glucuronoxylomannan (GXM), with or without rabbit anti-C. neoformans antiserum. At 3, 6, 12 and 24 h, AM were washed, lyzed and total RNA was isolated. Using reverse transcription-PCR, the transcripts of cytokine genes were semi-quantified by comparison with constitutive transcripts. Incubation of AM with lipopolysaccharide, as positive control, induced elevated levels of the three transcripts measured: interleukin (IL)-1 alpha, IL-6 and tumour necrosis factor (TNF)-alpha. Under the same conditions, no obvious changes were observed in the levels of transcription of these cytokines by AM after exposure to several strains of C. neoformans. However, AM that were incubated with both the yeast cells and rabbit polyclonal antisera to C. neoformans manifested significantly increased levels of mRNA for IL-6, but not IL-1 alpha or TNF-alpha. This increased level of IL-6 mRNA was detectable after incubation for 6 or 12 h. Levels of transcription in AM were unaffected by exposure to normal rabbit serum, specific antiserum alone. GXM at concentrations of 10, 100 or 500 micrograms ml-1, or GXM and antiserum. Adsorption of the antiserum with heat-killed yeast cells of C. neoformans diminished its ability to induce IL-6 mRNA in combination with fresh, viable yeast cells. The induction of IL-6 mRNA by yeast cells and antiserum does not require intact complement. In the absence of complement, the rabbit antiserum served as a potent opsonin and markedly increased phagocytosis of C. neoformans by AM. These results indicate that antibody-opsonized C. neoformans are readily phagocytosed by rat AM, and that antibody-mediated phagocytosis may differ from complement-mediated phagocytosis in the subsequent stimulation of IL-6.
收集Lewis大鼠肺泡巨噬细胞（AM）并在体外将其暴露于新型隐球菌，以研究炎性细胞因子的诱导情况。将组织培养孔中的AM与新型隐球菌的活酵母细胞或荚膜多糖葡糖醛酸木糖甘露聚糖（GXM）一起孵育，同时或不同时加入兔抗新型隐球菌抗血清。在3、6、12和24小时时，洗涤AM细胞，进行裂解并分离总RNA。使用逆转录聚合酶链反应（RT-PCR），通过与组成型转录本比较对细胞因子基因的转录本进行半定量分析。作为阳性对照，将AM与脂多糖一起孵育可诱导所检测的三种转录本水平升高：白细胞介素（IL）-1α、IL-6和肿瘤坏死因子（TNF）-α。在相同条件下，AM暴露于几种新型隐球菌菌株后，这些细胞因子的转录水平未观察到明显变化。然而，与酵母细胞和兔抗新型隐球菌多克隆抗血清一起孵育的AM，IL-6的mRNA水平显著升高，但IL-1α或TNF-α的mRNA水平未升高。孵育6或12小时后可检测到IL-6 mRNA水平的这种升高。AM中的转录水平不受暴露于正常兔血清或单独的特异性抗血清的影响。浓度为10、100或500微克/毫升的GXM，或GXM与抗血清。用新型隐球菌热灭活酵母细胞吸附抗血清会降低其与新鲜活酵母细胞联合诱导IL-6 mRNA的能力。酵母细胞和抗血清诱导IL-6 mRNA不需要完整的补体。在没有补体的情况下，兔抗血清作为一种有效的调理素，显著增加了AM对新型隐球菌的吞噬作用。这些结果表明，抗体调理的新型隐球菌很容易被大鼠AM吞噬，并且在随后刺激IL-6方面，抗体介导的吞噬作用可能与补体介导的吞噬作用不同。
