CXCR5 趨化激素受體調控樹突細胞移行之研究

Regulation of Dendritic Cells’ Migration by Chemokine Receptor CXCR5

吳孟澤(Meng-Ze Wu)

指導教授 ： 余幸司

高雄醫學大學/醫學院/醫學研究所/博士(2003年)

摘要

以往研究顯示, 樹突狀細胞（dendritic cells; 或簡稱 DC）所攜帶之特定趨化激素受體 (chemokine receptor)，在受到淋巴結內之趨化激素 (chemokine) 吸引後，能調控 DC 朝向淋巴結內特定的位置移動。 我們於本研究中發現, 由老鼠皮膚組織所分離出的遷移型樹突狀細胞（migratory DC; 或簡稱 migDC）可以大量表現趨化激素受體 CXCR5 mRNA（為原始朗格罕氏細胞 (Langerhans cells, 或簡稱 LC) 的50倍)，且經由體外試驗證實，此種遷移型樹突狀細胞會受到 B淋巴細胞趨化激素 (B lymphocyte chemoattactant, 或簡稱 BLC, 又稱 CXCL13) 的影響而產生趨化移動 (Chemotaxis)。當 migDC 由老鼠足部區注射進入體內後，約有 40% 比例的細胞會移動至鄰近淋巴結中的B淋巴細胞區 (B cell zones)。相對之下，另一種 DC ，起源於老鼠骨髓的 murine bone marrow-derived DC (或簡稱BMDC），其CXCR5 mRNA的表現量就只有 migDC 的十四分之一; 即使 in vitro 有 BLC 的存在下，也不會趨化移動; 而當BMDC 由老鼠足部區 (footpads) 注射進入體內後, 移動的標的主要侷限在淋巴結中的 T 淋巴細胞區 (T cell zones), 不會移動至淋巴結中的 B 淋巴細胞區。為了進一步釐清移動至B 淋巴細胞區的皮膚衍生型樹突狀細胞之起源 (來自表皮或真皮), 我們經由細胞表面抗原分析, 發現 13-35% 比例的皮膚衍生型 DC 能夠同時大量表現 MHC class II 分子與 CXCR5 蛋白，但值得注意的是，此種能夠大量表現 CXCR5 蛋白的皮膚衍生型 DC，並不具有一般表皮型 (epidermal type) DC 所擁有的 DEC205 分子, 據此可推測這些同時表現 MHC class II 分子與 CXCR5 蛋白的 DC 可能是來自皮膚真皮 (dermis) (而非表皮) 的樹突狀細胞. 此外，為了釐清 CXCR5 蛋白對於調控皮膚樹突狀細胞移行動向 (trafficking dynamics) 的重要性，本研究更藉由 retroviral gene transduction 的技術，將CXCR5 蛋白基因成功轉染 (transduction) 進入 BMDC 中。此經轉染的 BMDC (簡稱 CXCR5-BMDC), 表現出與野生型 BMDC 截然不同之表型: 不但能夠穩定表現CXCR5 蛋白，在 in vitro BLC 因子的存在下，也能產生趨化反應（3倍，p<0.01). 此外, 將此 CXCR5-BMDC 注射進入老鼠足部 (footpad), 約有40% 的 CXCR5-BMDC 會移動至淋巴結內的 B 細胞區域。為了進一步釐清 CXCR5 對 DC 移行動態之調控是否在抗原專一性之免疫反應的激發中扮演重要角色, 我們將經過 KLH (keyhole limpet hemocyanin) pulsed 之 CXCR5-BMDC 細胞或 vector-BMDC (只經過 retroviral vector 轉染之 BMDC, 做為對照組) 接種於鼠體並比較兩種細胞所引起之抗原特異性反應 (包含細胞性或體液性的免疫反應) 之不同，發現 CXCR5-BMDC 細胞所引起之足部腫脹程度 (footpad swelling) （遲延性過敏反應 (delayed-type hypersensitivity)之指標; 屬於細胞性免疫反應）較 vector-BMDC 細胞為小; 但卻會使老鼠產生較多之免疫球蛋白，IgG（p<0.05）與IgM（p<0.01）(體液性免疫反應)。我們因此推論活化型的皮膚樹突狀細胞會藉由自身 CXCR5 趨化激素受體蛋白的表現與外界 BLC 趨化激素的存在而受吸引移動至淋巴結中的 B 淋巴細胞區，並直接影響B淋巴球的活化。能夠表現 CXCR5 蛋白分子的樹突狀細胞應屬真皮衍生型，而 CXCR5 蛋白能夠導引 BMDC 細胞移行至淋巴結內的 B 細胞區域，並引發抗原專一性的免疫反應。

關鍵字

趨化激素 ； 趨化激素受體 ； 樹突細胞

並列摘要

Chemokine receptors on dendritic cells (DC) and chemokines within
lymph nodes (LN) contribute to trafficking of DC to appropriate sites
within the LN. Here we show that DC that have migrated out of skin ex
vivo (migratory DC, migDC) express 50-fold more CXCR5 mRNA than
fresh Langerhans cells and migrate in response to B lymphocyte
chemoattractant (BLC, or CXCL13) in vitro. When injected into the
footpad of mice, migDC emigrate to regional LN where up to 40 % are
found in B cell zones. By contrast, murine bone marrow-derived DC
display 14-fold less CXCR5, do not migrate to BLC in vitro, and migrate
strictly to T cell zones in LN. In order to determine the origin of
skin-derived migratory DC which are able to migrate to B cell zones of
draining lymph nodes, we analyzed the surface phenotype of skin-derived
migratory DC and found that 15–35% of MHC class II high cells showed
high levels of expression of CXCR5 but expressed low levels of DEC205, a
suggested characteristic of dermal-type DC in mice.

To study the effects of CXCR5 on the trafficking dynamics of DC, we
stably expressed CXCR5 in BMDC by retroviral gene transduction.
CXCR5 was detected by flow cytometry on transduced cells, which
responded to CXCL13 in vitro in chemotaxis assays (3-fold over
nontransduced BMDC, p < 0.01). When injected into the footpads of mice,
40% of injected CXCR5-BMDC were observed in BCZ of draining LN.
Mice were vaccinated with CXCR5- and vector-BMDC that were pulsed with keyhole limpet hemocyanin (KLH) to induce Ag-specific cellular and humoral immune responses. Mice injected with CXCR5-BMDC (vs
vector-BMDC) demonstrated marginally less footpad swelling in response to intradermal injection of KLH. Interestingly, significantly higher levels of
KLH-specific IgG (p < 0.05) and IgM (p < 0.01) were found in the serum of mice injected with CXCR5-BMDC compared with mice immunized with vector-transduced BMDC.
Thus, CXCR5 is predominantly expressed by dermal-type DC. Moreover, CXCR5 directs BMDC to BCZ of LN in vivo and modifies Ag-specific immune responses induced by BMDC vaccination. We propose that activated skin dermal-type DC utilize CXCR5 and BLC as a possible mechanism to home to B cell zones of LN, where they may have direct effects on B cells and help to induce antigen-specific humoral immune
responses.

並列關鍵字

chemokine ； chemokine receptor ； dendritic cell

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