Anti-CD45RB antibody deters xenograft rejection by modulating T cell priming and homing

Pancreatic islet xenotransplantation has been advocated as a way of overcoming the shortage of human donor tissue for the treatment of type 1 diabetes. However, the potent immune response against xenografts is a major barrier to their use. We show that a short course of the anti-CD45RB antibody, MB23G2, prolongs survival of fetal pig pancreas grafts in mice. To investigate this effect further we used an i.p. xenograft model in which both donor pig cells and host inflammatory cells can be expediently recovered and analyzed. Graft prolongation was associated with reduced T cell and macrophage infiltration, and reduced production of both T(h)1 and T(h)2 cytokines at the graft site. Graft survival was further increased and T cell infiltration further reduced by combining anti-CD45RB antibody with co-stimulation blockade. The primary effect of anti-CD45RB antibody may be on CD4 T cells, in keeping with the marked reduction in T cell cytokine production in both spleen and graft sites. This concurs with previous studies in allogeneic models that indicate that this antibody perturbs T cell responses by modifying signaling via the TCR. In addition, anti-CD45RB treatment led to reduced expression of LFA-1 and CD62 ligand (CD62L) on CD4 T cells, independent of antigenic challenge. LFA-1 may enhance co-stimulation, and both LFA-1 and CD62L are involved in T cell trafficking. Their reduced expression provides an explanation why the T cell pool is reduced in lymph nodes. We conclude that modulation of inflammation against xenografts by anti-CD45RB antibody is due to effects on both T cell priming and trafficking.     

转录因子IRF8抑制Th17细胞分化并减轻T细胞免疫介导的小鼠结肠炎

 目的：转录因子干扰素调节因子（interferon regulatory factor, IRF）家族与Th17的发育密切相关,近年来发现Th17细胞在炎症性肠病的发病中发挥重要作用,本研究探讨IRF8对Th17发育及T细胞转染免疫介导的小鼠实验性肠炎的影响。方法：(1)采用流式细胞术分选野生型（WT）或IRF8全基因敲除(IRF8 -/-)小鼠脾脏和淋巴结的naive CD4 +T细胞(CD4 + CD62L +CD44 low),在Th1、Th2或Th17极化的条件下培养,采用流式细胞术检测Th1、Th2和Th17的比例。(2)建立实验性肠炎模型：采用免疫磁珠法分选WT或IRF8 -/-小鼠中的脾脏和淋巴结中CD4 +CD25 +Treg,WT小鼠的CD4 + CD45RB hi T细胞单独或者分别联合WT或IRF8 -/-小鼠的CD4 +CD25 +Treg腹腔注射给RAG1 -/-小鼠;WT或IRF8 -/-小鼠的naive CD4 + CD45RB hi T细胞腹腔注射给RAG1 -/-小鼠;观察上述小鼠每周体重的变化,第5周时处死小鼠,进行结肠炎病理评分和肠系膜淋巴结T淋巴细胞亚群检测。结果：(1)IRF8 -/-较WT的naive CD4 +T细胞在极化条件下向Th17细胞分化更明显(P<001),而对Th1和Th2细胞的分化无影响(P>0.05)。(2)CD4 + CD45RB hi T细胞转染给RAG1 -/-小鼠,IRF8 -/-较WT供体鼠引起的RAG1 -/-小鼠体重显著降低(P<0.05),结肠炎评分显著增高（P<0.05）,且肠系膜淋巴结中IL-17 +CD4 +细胞比例明显增高(P<0.01),而 IFN-γ +CD4 + 和 Foxp3 +CD4 +细胞比例无影响(P>0.05);IRF8 -/-小鼠的CD4 +CD25 +Treg对WT小鼠CD4 + CD45RB hi T细胞转染给RAG1 -/-小鼠诱发的免疫介导的结肠炎显示出正常的免疫抑制作用。结论：转录因子IRF8基因敲除促进CD4 +T细胞向Th17细胞分化,促进转染naive CD4 +T细胞诱导的实验性结肠炎的发生,IRF8基因敲除小鼠Treg细胞免疫抑制功能正常。     

IL-2协同抗CD45RB抗体调节Treg/Th17细胞的比例并延长小鼠移植皮肤存活时间

目的:研究IL-2在抗CD45RB抗体诱导免疫耐受中对Treg/Th17细胞分化的影响,进一步阐明抗CD45RB抗体诱导免疫耐受的机制。方法:用免疫磁珠分选C57BL/6小鼠脾脏中的CD4+T细胞,在抗CD45RB抗体与IL-2的作用下培养72小时后,流式检测Treg/Th17细胞的变化。以BALB/c小鼠为供体,C57BL/6小鼠为受体建立同种异基因皮肤移植模型,分别给予抗CD45RB抗体及IL-2等治疗,术后1、3、5、7、9天取受体鼠脾细胞,动态检测Treg/Th17细胞的变化;术后第9天取移植皮肤HE染色观察炎性细胞的浸润情况;观察并记录移植皮肤的存活时间。结果:CD4+T细胞在IL-2联合抗CD45RB抗体的作用下培养72小时后,Treg比例升高,Th17细胞比例下降;IL-2联合抗CD45RB抗体治疗后明显延长小鼠移植皮肤的存活时间。结论:IL-2可以明显增强抗CD45RB抗体诱导免疫耐受的形成,使Treg细胞上调,下调Th17细胞,有利于免疫耐受的形成。     

alpha4 integrins and L-selectin differently orchestrate T-cell activity during diabetes prevention following oral administration of CTB-insulin

Oral administration of insulin conjugated to the B chain of cholera toxin (CTB-insulin) in non-obese diabetic (NOD) mice results in diabetes prevention. We investigated the respective contributions of L-selectin (CD62L) and alpha4-integrin pathways during CTB-driven tolerance. Purified CD62L+CD4+ cells from CTB-insulin fed mice significantly reduced the capacity of diabetogenic T cells to transfer diabetes in syngeneic recipients. In vivo antibody blockade of fed animals during adoptive co-transfer experiments indicated that both CD62L and alpha4-integrins pathways were necessary to develop a protective response after oral tolerance induction. In contrast, when antibodies were given to recipient mice, only CD62L was critical for the protection. In vitro stimulated CD62L+CD4+ cells from the spleen of fed animals secreted lower amounts of IL-4 and IL-10 but comparable levels of TGFbeta than CD62L-cells. A reduced IFN-gamma production between the two cell subsets was specifically observed in CTB-insulin fed mice. Furthermore, antibody treatments induced changes in T-cell migration to the spleen, mesenteric and pancreatic lymph nodes. The protective effect was also associated with migration of regulatory T cells into pancreatic islets. Taken together, our results suggest that L-selectin and alpha4-integrin have distinct but complementary roles in the generation and function of regulatory CD4+ T cells following CTB-insulin administration.     

Inducible costimulator controls migration of T cells to the lungs via down-regulation of CCR7 and CD62L

We and others reported that inducible costimulator-deficient (ICOS(-/-)) mice manifest a defect in Th2-mediated airway inflammation, which was attributed to reduced Th2 differentiation in the absence of ICOS signaling. Interestingly, the number of CD4 T cells present in the airways and lungs after sensitization and challenge is significantly reduced in ICOS(-/-) mice. We now show that this reduction is not attributable simply to a reduced proliferation of ICOS(-/-) cells, because significantly more ICOS(-/-) than wild-type activated CD4 T cells are present in the lymph nodes, suggesting that more ICOS(-/-) CD4 T cells than wild-type CD4 T cells migrated into the lymph nodes. Further investigation revealed that activated ICOS(-/-) CD4 T cells express higher concentrations of the lymph node homing receptors, CCR7 and CD62L, than do wild-type CD4 T cells, leading to a preferential return of ICOS(-/-) cells to the nondraining lymph nodes rather than the lungs. Blocking reentry into the lymph nodes after the initiation of Th2-mediated airway inflammation equalized the levels of CD4 and granulocyte infiltration in the lungs of wild-type and ICOS(-/-) mice. Our results demonstrate that in wild-type CD4 T cells, co-stimulation with ICOS promotes the down-regulation of CCR7 and CD62L after activation, leading to a reduced return of activated CD4 T cells to the lymph nodes and a more efficient entry into the lungs.     

FTY720, a novel immunosuppressant, induces sequestration of circulating mature lymphocytes by acceleration of lymphocyte homing in rats, III. Increase in frequency of CD62L-positive T cells in Peyer's patches by FTY720-induced lymphocyte homing.

FTY720, a novel immunosuppressant, sequesters circulating mature lymphocytes, especially T cells, within lymph nodes and Peyer's patches by accelerating lymphocyte homing, and thereby causes lymphocyte depletion in the blood. The FTY720-induced acceleration of lymphocyte homing appears to be mediated by lymphocyte homing receptors including CD62L, CD49d/beta7, and CD11a/CD18. In this study, expressions of CD62L, CD49d and CD11a on T cells in the peripheral blood, lymph nodes and Peyer's patches were analysed by flow cytometry in rats given FTY720 (1 mg/kg) orally. FTY720 markedly decreased the number of peripheral blood T cells, while not affecting CD62L, CD49d and CD11a expressions at 1-3 hr after administration. In contrast, both the frequency of CD62L-positive T cells and intensity of CD62L expression on T cells were increased in Peyer's patches but not lymph nodes at 3 hr after administration of FTY720. CD49d and CD11a expressions on T cells were unaffected by FTY720 in both Peyer's patches and lymph nodes at the same point in time. On the other hand, analysis of lymphocyte homing with calcein-labelled lymphocytes and anti-CD62L monoclonal antibody (mAb) confirmed that FTY720 predominantly increased CD62L-dependent lymphocyte homing to Peyer's patches. These findings indicate that FTY720 increases the frequency of CD62L-positive T cells by accelerating CD62L-predominant homing in Peyer's patches.     

Dynamics of CD62L/CD45RB CD4+ and CD8+ lymphocyte subsets in hepatic and splenic tissues during murine visceral leishmaniasis

This study aimed to characterise, for the first time, the dynamics of CD4+ and CD8+ lymphocyte CD62L/CD45RB subsets, during visceral leishmaniasis. Memory/activated status of hepatic and splenic T cells was compared in mice strains with "cure" and "non-cure" phenotypes to Leishmania infantum infection. In both mice strains, a correlation between the dynamics of the memory CD4+ and CD8+ T cells (CD62Llow/CD45RBlow) subsets in the liver and the pre-activated phenotype of lymphocytes (CD62Llow/CD45RBhigh) from the spleen was detected suggesting that this organ is the source of Leishmania-specific T lymphocytes that migrate to the liver, where parasite replication is highly active. In the liver, these pre-activated cells become effector T lymphocytes, however, a strong regulation of CD8+ T cell effector function was observed, probably preventing hepatic tissue damage. Comparing mice strains with "cure" and "non-cure" phenotype, an imbalance between "protective" CD45RBhigh and "pathogenic" CD45RBlow CD4+ subsets in B10.D2/n animals might be involved in the evolution of a non-healing infection.     

B淋巴细胞在抗CD45RB抗体诱导的移植免疫耐受中的作用

 目的: 探讨B淋巴细胞在抗CD45RB抗体诱导的移植免疫耐受中的作用。方法: 抗CD45RB抗体对BALB/c裸鼠进行预处理后制备脾脏单细胞悬液,与BALB/c小鼠T淋巴细胞和C57BL/6小鼠脾细胞混合培养,流式细胞术分析Th1、Th2、Treg和Tm淋巴细胞。以B6.μMT^-/-小鼠为受体、BALB/c小鼠为供体建立皮肤移植模型,移植后向受体鼠腹腔注射抗CD45RB单抗,监测脾淋巴细胞CD3⁺CD45RB^hi细胞比例。在混合淋巴培养过程中加入抗CD45RB单抗,分离B细胞,建立以BALB/c小鼠为供体、B6.μMT^-/-小鼠为受体的心脏移植模型,通过尾静脉注射B细胞给B6.μMT^-/-小鼠,观察受体鼠生存期和B细胞分布。结果: 在裸鼠体内用抗CD45RB抗体处理过的B淋巴细胞,与T淋巴细胞混合培养时,可使Treg和Th2淋巴细胞比例明显升高,Th1淋巴细胞的比例明显下降,Tm细胞无明显变化。在体内B淋巴细胞缺失的情况下,抗CD45RB抗体依然能够降低T细胞表面CD45RB的表达,与对照组B淋巴细胞存在组相比,抗CD45RB抗体对T淋巴细胞表面CD45RB下调更为快速,但最终CD3⁺CD45RB^hi T细胞比例无明显变化。体外抗CD45RB抗体处理过的B淋巴细胞可以延长受体鼠的生存时间。B6.μMT^-/-鼠在接受抗CD45RB抗体处理的B细胞并进行同种异体心脏移植后,B细胞可向胸腺迁移。结论: 在抗CD45RB抗体诱导的免疫耐受中,B淋巴细胞可能通过介导各T淋巴细胞亚群比例发挥着重要作用,且在中枢耐受中也起到一定作用,但是仅靠B淋巴细胞无法形成完全耐受。     

Occurrence of interleukin-5 production by CD4- CD8- (double-negative) T cells in lungs of both normal and congenitally athymic nude mice infected with Toxocara canis.

We studied cells in the lungs of BALB/c and BALB/c-nu/nu (nude) mice infected with Toxocara canis, which produced interleukin-5 (IL-5) in in vitro culture with larval excretory-secretory antigen (ESAg). The proportion of CD4+/CD8+/CD4- CD8- cells in lungs of both BALB/c and nude mice was unchanged before and after infection with T. canis. Panning and complement-mediated lysis using monoclonal antibody (mAb) to CD4 showed that CD4+ cells in the lung from both mice produced IL-5. Anti-CD4 mAb suppressed ESAg-stimulated IL-5 production in vitro. In vitro depletion or inhibition of CD8+ cells reduced IL-5 production significantly in some cases, suggesting involvement with IL-5 production. Anti-CD3 mAb enhanced IL-5 production when incubated with or without ESAg. Production of IL-5 was reduced by in vivo depletion of CD4+ cells only and both CD4+ and CD8+ T cells, by intraperitoneal injection with appropriate mAb; IL-5 production was stimulated by anti-CD3 mAb. In contrast, IL-5 production by lung cells of BALB/c mice decreased by more than 90% after simultaneous injection with anti-CD4, anti-CD8 and anti-CD3 mAb, and was not enhanced by anti-CD3 mAb. Similar results were obtained in nude mice. These results suggest that CD4- CD8- T cells, as well as CD4+ T cells, produce IL-5.     

Apoptosis induced in HIV-1-exposed, resting CD4+ T cells subsequent to signaling through homing receptors is Fas/Fas ligand-mediated

The hallmark of HIV-1 disease is the gradual disappearance of CD4+ T cells from the blood. The mechanism of this depletion, however, is still unclear. Evidence suggests that lymphocytes die in lymph nodes, not in blood, and that uninfected bystander cells are the predominant cells dying. Our and others' previous studies showed that the lymph node homing receptor, CD62 ligand (CD62L), and Fas are up-regulated on resting CD4+ T cells after HIV-1 binding and that these cells home to lymph nodes at an enhanced rate. During the homing process, signals are induced through various homing receptors, which in turn, induced many of the cells to undergo apoptosis after they entered the lymph nodes. The purpose of this study was to determine how the homing process induces apoptosis in HIV-1-exposed, resting CD4+ T cells. We found that signaling through CD62L up-regulated FasL. This resulted in apoptosis of only HIV-1-presignaled, resting CD4+ T cells, not normal CD4+ T cells. This homing receptor-induced apoptosis could be blocked by anti-FasL antibodies or soluble Fas, demonstrating that the Fas-FasL interaction caused the apoptotic event.     

CD4~+CD25~+CD127~-T细胞在抗CD45RB抗体诱导免疫耐受中的作用

目的:研究CD4+CD25+CD127-T细胞在抗CD45RB抗体诱导的免疫耐受中所发挥的作用,从而阐明抗CD45RB抗体诱导免疫耐受作用的机制。方法:体内实验建立小鼠异位心脏移植模型,观察抗CD45RB抗体对移植物生存期的影响。体外实验观察抗CD45RB抗体对T细胞增殖抑制能力和对CD4+CD25+CD127-T细胞生成的影响。流式细胞术检测外周血和混合细胞中CD4+CD25+CD127-T细胞百分率,ELISA法检测血清和培养液中IL-2和IL-10含量,Real-TimePCR法检测脾脏和混合细胞中Foxp3基因的表达,移植心脏病理学观察。结果:抗CD45RB抗体显著延长移植物存活时间(P0.01),对ConA刺激引起的T淋巴细胞增殖具有明显的抑制能力(P0.05)。与对照组相比,实验组CD4+CD25+CD127-T细胞百分率和Foxp3 mRNA表达量均明显增加(P0.05),实验组IL-2水平较对照组明显降低(P0.05),但IL-10含量较对照组明显升高(P0.05)。病理结果显示对照组移植心脏出现典型细胞免疫性损伤病理改变,而实验组中几乎无炎性细胞浸润现象。结论:抗CD45RB抗体能显著延长移植物存活时间,其诱导免疫耐受机制与上调CD4+CD25+CD127-T细胞百分率和增加Foxp3 mRNA表达量有关。     

CD62L is required for the priming of encephalitogenic T cells but does not play a major role in the effector phase of experimental autoimmune encephalomyelitis

CD62L (l ‐selectin, mel 14) regulates naïve T cell homing into lymph nodes and the migration of leucocytes to sites of inflammation. The requirement of CD62L in the pathogenesis of experimental autoimmune encephalomyelitis (EAE), an animal model of multiple sclerosis, has been demonstrated previously. However, it remains controversial as to whether CD62L is required for the induction or the effector phase of EAE. It is also unclear whether other non‐T effector cells need CD62L to enter the central nervous system (CNS) parenchyma and exert their damaging effects on myelin. We report that mice with a targeted mutation of CD62L are resistant to Myelin oligodendrocyte glycoprotein peptide‐induced EAE. CD62L‐deficient mice had no peptide‐specific T cell responses in the draining lymph nodes and had lower levels of peptide‐specific T cell responses in spleens at a later time point. Adoptive transfer studies showed that CD62L‐deficient mice were fully susceptible to adoptive transfer EAE induced by either wildtype or CD62L‐deficient T cells. Moreover, CD62L‐deficient, F4/80⁺ macrophages can be efficiently recruited into the CNS parenchyma. These data suggest that CD62L is required for the induction of encephalitogenic T cells during EAE development, but is not required by T and non‐T effector cells to attack the CNS parenchyma.     

Marked increase in L-selectin-negative T cells in neonatal pertussis. The lymphocytosis explained?

The detailed immunophenotype of peripheral blood lymphocytes from a neonate with pertussis was determined by flow cytometry and compared with results from cord blood from healthy newborns. Most (72%) of the lymphocytes were CD3+ T cells with a normal CD4/CD8 ratio (2.5). The T cells were largely HLA-DR negative and CD45RA+, consistent with unstimulated na?ve T cells. Almost all of the CD4+ T cells were Leu8 (L-selectin, CD62L) negative, while almost all of the CD8+ T cells were CD28+. There was no increase in CD7- CD4+ T cells (Th2-like). No relative increase in CD16/56+ NK cells (5%) or CD19/20+ B cells was seen. The most dramatic finding in this case was the remarkable lack of expression of L-selectin by the T cells. L-selectin expression is associated with homing of peripheral blood lymphocytes to lymph nodes. The dramatic reduction in L-selectin expression of the T lymphocytes in pertussis, perhaps induced by pertussis toxin, likely prevents homing of the T cells to peripheral lymphoid tissues and provides a likely explanation for the marked lymphocytosis noted in this disease.     

Role of L-selectin in the development of autoimmune diabetes in non-obese diabetic mice

Autoimmune diabetes is characterized by an early mononuclear infiltration of pancreatic islets and later selective autoimmune destruction of insulin-producing beta cells. Lymphocyte homing receptors have been considered candidate targets to prevent autoimmune diabetes. L-selectin (CD62L) is an adhesion molecule highly expressed in naive T and B cells. It has been reported that blocking L-selectin in vivo with a specific antibody (Mel-14) partially impairs insulitis and diabetes in autoimmune diabetes-prone non-obese diabetic (NOD) mice. In the present study we aimed to elucidate whether genetic blockade of leukocyte homing into peripheral lymph nodes would prevent the development of diabetes. We backcrossed L-selectin-deficient mice onto the NOD genetic background. Surprisingly NOD/L-selectin-deficient mice exhibited unaltered islet mononuclear infiltration, timing of diabetes onset and cumulative incidence of spontaneous diabetes when compared to L-selectin-sufficient animals. CD4, CD8 T cells and B cells were present in islet infiltrates from 9-week-old L-selectin-sufficient and -deficient littermates. Moreover, total splenocytes from wild-type, heterozygous or NOD/L-selectin-deficient donor mice showed similar capability to adoptively transfer diabetes into NOD/SCID recipients. On the other hand, homing of activated, cloned insulin-specific autoaggressive CD8 T cells (TGNFC8 clone) is not affected in NOD/L-selectin-deficient recipients. We conclude that L-selectin plays a small role in the homing of autoreactive lymphocytes to regional (pancreatic) lymph nodes in NOD mice.     

抗原特异性初始CD4+T细胞的体内分化及特性

为了探讨抗原特异性CD4+T细胞在体内的分裂、表型、Th1细胞因子的产生和组织器官的分布。将CFSE标记的抗原特异性初始CD4+T细胞静脉被动输给小鼠后,进行免疫,3d后处死小鼠取其脾脏、淋巴结和肺组织,分离单个核细胞,利用流式细胞计数仪在单个细胞水平上,观察细胞的分裂、表型、Th1细胞因子的产生和组织分布。结果显示在没有抗原刺激的情况下,未见初始CD4+T细胞分裂,其主要分布于淋巴结和脾脏。当受到抗原刺激后,CD4+T细胞分裂1～5次,主要分布于脾脏和肺组织,CD25的表达增加,CD62L的表达随着细胞分裂次数的增加而减少。IL-12促进CD25的表达和细胞的分裂。促进Th1细胞的分化和IFN-γ的表达。研究的结果提示,在体内,当CD4+T细胞活化后,主要分布于脾和非淋巴组织发挥其免疫效应。     

The phenotype and survival of antigen-stimulated transgenic CD4 T cells in vivo: the influence of persisting antigen

Naive and primed/memory CD4 T cells are distinguished by changes in the expression of activation/adhesion molecules that correspond with an altered function. Adoptively transferred TCR transgenic (tg) CD4 T cells specific for ovalbumin peptide (OVA-pep) were analysed for changing phenotype and the speed of change in vivo following antigen challenge with alum-precipitated (ap) OVA-pep, a conjugate that stimulated a Th2-type cytokine response. The change of CD45RB in relation to number of divisions showed that the transition from CD45RB(hi) (naive) to CD45RB(low) (primed/memory) was incremental; with each cell cycle the number of CD45RB(hi) molecules on the cell surface was diluted by approximately half and replaced by the low-weight isoform. Similarly, the change to CD44(hi) expression increased gradually during four rounds of proliferation. The loss of CD62L expression occurred early and was independent of cell division. CD69 was up-regulated quickly within 1-2 cycles, but down-regulated after about seven divisions. The expression of CD49d was not altered during the early rounds of division, although it was up-regulated on 30-60% of tg T cells dividing repeatedly (>or=8 cycles). When analysed on day 3 following stimulation, CD25 was no longer up-regulated. The intra-peritoneal injection of ap-OVA-pep stimulated tg T cells in the spleen and mesenteric lymph node one day in advance of those in more distant peripheral lymph nodes. Evidence indicated that residual antigen persisted for at least 4 weeks and was able to stimulate naive tg T cells. However, residual antigen had no net effect on extending or reducing survival of the transferred population.     

Bcl6 is essential for the generation of long-term memory CD4+ T cells

Bcl6 plays a role in the generation and maintenance of memory CD8(+) T cells. We analyzed here a role for Bcl6 in the generation of long-term memory CD4(+) T cells. Naive CD45RB(+) CD4(+) T cells from Bcl6-deficient DO11.10 (KJ1.26(+)) transgenic mice were transferred into BALB/c mice and immunized with ovalbumin peptide and LPS. Long-term memory KJ1.26(+) CD4(+) T cells from wild-type mice were detected in the spleen, lungs and liver during 10 weeks after immunization; however, Bcl6-deficient KJ1.26(+) CD4(+) T cells were vanished completely in those organs 4 weeks after immunization. Since memory CD4(+) T cells can be generated from effector CD4(+) T cells, properties of Bcl6-deficient effector CD4(+) T cells were compared with those wild-type effector CD4(+) T cells 10 days after immunization. Numbers of IFN-gamma-non-producing CD45RB(-), CD62L(+) or IL-7Ralpha(+) effector CD4(+) T cells in the spleen, lungs and liver were similar between Bcl6-deficient and wild-type CD4(+) T cells. However, the percentage of apoptotic cells in Bcl6-deficient effector CD4(+) T cells was higher than that in wild-type effector CD4(+) T cells. At the late effector phase, the number of IFN-gamma-non-producing cells and the percentage of apoptotic cells in Bcl6-deficient CD4(+) T cells were smaller and higher than those in wild-type CD4(+) T cells, respectively. These data suggest that Bcl6 in CD4(+) T cells plays a role in protection of memory precursor CD4(+) T cells from apoptosis and may involve in survivability of long-term memory CD4(+) T cells.     

Distinct compartmentalization of CD4+ T-cell effector function versus proliferative capacity during pulmonary cryptococcosis

The activation and expansion of T cells and their acquisition of effector function are key steps in the development of the adaptive immune response. Most infections are predominantly outside of the lymphoid tissues, and it is unclear at what point developmentally and anatomically T cells acquire effector function in vivo. In these studies, we compared the activation and polarization of T cells during murine pulmonary Cryptococcus neoformans infection in the secondary lymphoid tissues and at the site of primary infection. Few CD4(+) and CD8(+) T cells expressed an activated phenotype (CD44(hi,) CD25(+), CD69(+), CD62L(lo), CD45RB(lo)) at the sites of clonal expansion (lymph nodes, spleen, and blood). In contrast, a high percentage of T cells expressed activation markers at the site of primary infection, the lungs. Additionally, the polarization of CD4(+) T cells to interferon-gamma-producing effector cells occurred at the site of infection, the lungs. CD4(+) and CD8(+) T cells from secondary lymphoid organs responded to TCR restimulation by proliferating, whereas T cells from the lungs proliferated poorly. This report demonstrates for the first time that T-cell activation and effector function in secondary lymphoid tissues during fungal infection is characteristically different from that at the site of primary infection.     

Novel phenotypes and migratory properties distinguish memory CD4 T cell subsets in lymphoid and lung tissue

Memory T cells are heterogeneous in expression of lymph node homing receptors, delineating "central-memory" (TCM, CD62Lhi/CCR7+) and "effector-memory" (TEM, CD62Llo/CCR7-) subsets that migrate to lymphoid and non-lymphoid tissues, respectively. It is not known how these subsets arise or how homing receptor expression and tissue origin determine their functional and migratory properties. Here, we investigated the role of CD62L expression in the generation, function, distribution and migration of heterogeneous memory CD4 T cells specific for influenza hemagglutinin (HA). We found that CD62Lhi and CD62Llo memory subsets are generated independent of CD62L expression by the activated precursor, and both subsets distribute into spleen and lung. Functionally, spleen- and lung-derived CD62L memory subsets produce effector cytokines at similar kinetics but differ strikingly in cell surface phenotype and migration: the CD62Llo memory subset expresses a classic memory phenotype (CD45RBlo/CD44hi/CD11a(hi)), while the CD62Lhi subset expresses an unconventional phenotype (CD45RBhi/CD44int/CD11a(int)), defining a new polyclonal memory subset. The CD62Lhi subset also trafficked more efficiently than CD62Llo cells into lymph nodes; however, only lung but not spleen CD62Llo memory T cells homed to lung. Our results reveal novel phenotypic heterogeneity of memory CD4 T cells co-segregating with CD62L expression and tissue-specific tropism of non-lymphoid memory CD4 T cells.     

On the role of CD26 in CD4 memory T cells

We studied an in vivo mouse model to evaluate the relationships between CD26--a glycoprotein with dipeptidyl peptidase IV (DPP-IV) activity implicated in the regulation of immune functions--and T cells expressing the effector/memory phenotype CD45RB. We report that CD26 does not define a differentiation stage of CD4 T cells because the density and frequency of CD26 on CD4 T cells from the spleen, inguinal and mesenteric lymph node was similar within the CD45RB+ (na?ve) and CD45RB- (antigen primed) subsets. This observation was confirmed using CD4 T cells from a T-cell receptor transgenic (tg) model. CD4 tg T cells specific for ovalbumin (OVA) were adoptively transferred and challenged in vivo with antigen. CD26 expression was the same on naive and antigen-stimulated CD4 T cells. Depleting CD4 T cells with an anti-CD4 antibody preferentially depleted the CD45RB+ subset. In CD4 depleted animals CD26 expression was not altered on the CD45RB- subset but the density of CD26 was marginally increased on the remaining CD45RB+ CD4 T cells. The results suggest that, unlike the human, CD26 in the mouse was not directly linked with T cell activation.