High Density Insulin Receptor-Positive Diabetogenic Tlymphocytes in Nonobese Diabetic Mice are Memory Cells

Abstract

Our previous work examining the importance of insulin receptor (IR) expression on T cells has demonstrated that when T cells from nonobese diabetic mice were sorted into populations expressing a high (IR^High) and a low (IR^Low) density of IR, IR^High T cells rapidly transferred insulitis and diabetes. We have further characterized IR^High T cells. Both CD4⁺ and CD8⁺ cells were detected in the IR^High T cell population, but IR^High expression was detected predominantly on CD4⁺ cells. IR^High T cells were polyclonal for TCR Vβ-chain expression. By 3 color flow cytometric analysis, virtually all IR^High T cells expressed low or negligible levels of CD62L (CD62L^Low/-) and high levels of CD44 (CD44^High). The lack of IL-2 receptor and transferrin receptor expression as seen previously, together with the CD62L^Low/- CD44^High phenotype suggests that IR^High T cells are memory cells. However, since only about one quarter of all of the CD62L^Low/- or CD44^High T cells were also IR^High, the IR^High phenotype defines a subpopulation of memory T cells that are aggressively diabetogenic.     

Defective induction of the proteasome associated with T‐cell receptor signaling underlies T‐cell senescence

The proteasome degradation machinery is essential for a variety of cellular processes including senescence and T‐cell immunity. Decreased proteasome activity is associated with the aging process; however, the regulation of the proteasome in CD4⁺ T cells in relation to aging is unclear. Here, we show that defects in the induction of the proteasome in CD4⁺ T cells upon T‐cell receptor (TCR) stimulation underlie T‐cell senescence. Proteasome dysfunction promotes senescence‐associated phenotypes, including defective proliferation, cytokine production and increased levels of PD‐1⁺ CD44^High CD4⁺ T cells. Proteasome induction by TCR signaling via MEK‐, IKK‐ and calcineurin‐dependent pathways is attenuated with age and decreased in PD‐1⁺ CD44^High CD4⁺ T cells, the proportion of which increases with age. Our results indicate that defective induction of the proteasome is a hallmark of CD4⁺ T‐cell senescence.     

CD146 Expression Influences Periapical Cyst Mesenchymal Stem Cell Properties

Recent studies have identified a new human dental derived progenitor cell population with multi-lineage differentiation potential referred to as human periapical cyst mesenchymal stem cells (hPCy-MSCs). In the present study, we compared two subpopulations of hPCy-MSCs characterised by the low or high expression of CD146 to establish whether this expression can regulate their stem cell properties. Using flow cytometry, we evaluated the stem cell marker profile of hPCy-MSCs during passaging. Furthermore, CD146^Low and CD146^High cells were sorted by magnetic beads and subsequently both cell populations were evaluated for differences in their proliferation, self-renewal, stem cell surface markers, stemness genes expression and osteogenic differentiation potential.We found that hPCy-MSCs possessed a stable expression of several mesenchymal stem cell surface markers, whereas CD146 expression declined during passaging.In addition, sorted CD146^Low cells proliferated significantly faster, displayed higher colony-forming unit-fibroblast capacity and showed higher expression of Klf4 when compared to the CD146^High subset. Significantly, the osteogenic potential of hPCy-MSCs was greater in the CD146^Low than in CD146^High population. These results demonstrate that CD146 is spontaneously downregulated with passaging at both mRNA and protein levels and that the high expression of CD146 reduces the proliferative, self-renewal and osteogenic differentiation potential of hPCy-MSCs. In conclusion, our study demonstrates that changes in the expression of CD146 can influence the stem cell properties of hPCy-MSCs.     

Methoxyamine sensitizes the resistant glioblastoma T98G cell line to the alkylating agent temozolomide

Chemoresistance represents a major obstacle to successful treatment for malignant glioma with temozolomide. N ⁷-methyl-G and N ³-methyl-A adducts comprise more than 80 % of DNA lesions induced by temozolomide and are processed by the base excision repair, suggesting that the cellular resistance could be caused, in part, by this efficient repair pathway, although few studies have focused on this subject. The aim of this study was to evaluate the cellular responses to temozolomide treatment associated with methoxyamine (blocker of base excision repair) in glioblastoma cell lines, in order to test the hypothesis that the blockage of base excision repair pathway might sensitize glioblastoma cells to temozolomide. For all the tested cell lines, only T98G showed significant differences between temozolomide and temozolomide plus methoxyamine treatment, observed by reduced survival rates, enhanced the levels of DNA damage, and induced an arrest at G2-phase. In addition, ~10 % of apoptotic cells (sub-G1 fraction) were observed at 48 h. Western blot analysis demonstrated that APE1 and FEN1 presented a slightly reduced expression levels under the combined treatment, probably due to AP sites blockade by methoxyamine, thus causing a minor requirement of base excision repair pathway downstream to the AP removal by APE1. On the other hand, PCNA expression in temozolomide plus methoxyamine-treated cells does not rule out the possibility that such alteration might be related to the blockage of cell cycle (G2-phase), as observed at 24 h of recovery time. The results obtained in the present study demonstrated the efficiency of methoxyamine to overcome glioblastoma resistance to temozolomide treatment.     

Ag-specific type 1 CD8 effector cells enhance methotrexate-mediated antitumor responses by modulating differentiated T cell localization, activation and chemokine production in established breast cancer

The chemotherapeutic agent methotrexate is widely used in the treatment of breast cancer. Although its mechanism-of-action has been defined, less is known about its interaction with T cell-mediated antitumor responses. Type 1 CD8 T cell-mediated immune responses (Tc1) are cytolytic, produce IFN-gamma and are associated with effective antitumor responses. Using a murine transgenic TCR tumor model, we show that single-dose treatment with methotrexate enhanced CD8-mediated type 1 antitumor responses when administered 3 days prior to Tc1 effector cell transfer. Co-treatment with methotrexate not only enhanced donor Tc1 cell accumulation and persistence at sites of primary tumor growth, but also promoted elevated levels of activated donor TIL cells. This markedly enhanced the appearance of endogenous differentiated (CD44^High) CD8 tumor-infiltrating cells when compared to that of corresponding groups receiving either MTX or Tc1 cell transfer alone. Such cells were acutely activated as defined by co-expression of surface markers associated with TCR engagement (CD69) and T cell activation (CD25) at both early (days 1–8) and late (days 12–20) stages following treatment. Conversely, such animals showed an early decrease in CD4⁺/CD44^High/CD25⁺/CD69⁺ T cells that correlated with delays in tumor growth in vivo. Moreover, cellular response kinetics appeared to further correlate with the up-regulation of endogenous T cells producing the chemokine IP-10 in vivo. This suggested that Tc1 cell transfer, in combination with chemotherapy, can enhance antitumor responses by modulating immunoregulatory T cells involved in homeostasis and immune tolerance within the tumor environment. These studies offer insight into mechanisms that enhance T cell-based immunotherapy in cancer.     

Ag-Specific Type 1 CD8 Effector Cells Enhance Methotrexate-Mediated Antitumor Responses by Modulating Endogenous CD49b-Expressing CD4 and CD8 T Effector Cell Subpopulations Producing IL-10

The chemotherapeutic agent methotrexate is widely used in the treatment of breast cancer. Although its mechanism-of-action has been defined, less is known about its interaction with Ag-specific T cell-mediated antitumor responses. Type 1 CD8 T cell-mediated immune responses (Tc1) are cytolytic, produce IFN-gamma and are associated with effective antitumor responses. Using a murine transgenic TCR tumor model, we show that single-dose-treatment with methotrexate enhanced CD8-mediated type 1 antitumor responses when administered three days prior to Tc1 effector cell transfer. Co-treatment with methotrexate not only enhanced donor Tc1 cell accumulation and persistence at sites of primary tumor growth, but also promoted elevated levels of activated CD25⁺ expressing donor TIL cells. This correlated with a marked decrease in the appearance of endogenous differentiated (CD44^High) CD3/CD8/CD49b and CD3/CD4/CD49b tumor-infiltrating effector T cells at both early (Days 1–8) and late (Days 12–20) stages following treatment when compared to that of corresponding groups receiving either MTX or Tc1 cell transfer alone. Moreover, such cellular response kinetics appeared to further correlate with the down-regulation of endogenous CD4/CD44^High/CD49b effector T cells producing IL-10 and delays in tumor growth in vivo. This suggested that Ag-specific Tc1 cell transfer, in combination with chemotherapy, can enhance antitumor responses by modulating select CD49b-expressing T effector/memory cell subpopulations involved in homeostasis and immune tolerance within the tumor environment. These studies offer insight into mechanisms that enhance T cell-based immunotherapy in cancer. Supplementary materials are available for this article. Go to the publisher's online edition of Immunological Investigations for the following free supplemental resource(s): Addendum 1.     

Mesenteric lymph node CD11b− CD103+ PD‐L1High dendritic cells highly induce regulatory T cells

Dendritic cells (DCs) in mesenteric lymph nodes (MLNs) induce Foxp3⁺ regulatory T cells to regulate immune responses to beneficial or non‐harmful agents in the intestine, such as commensal bacteria and foods. Several studies in MLN DCs have revealed that the CD103⁺ DC subset highly induces regulatory T cells, and another study has reported that MLN DCs from programmed death ligand 1 (PD‐L1) ‐deficient mice could not induce regulatory T cells. Hence, the present study investigated the expression of these molecules on MLN CD11c⁺ cells. Four distinct subsets expressing CD103 and/or PD‐L1 were identified, namely CD11b⁺ CD103⁺ PD‐L1^High, CD11b⁻ CD103⁺ PD‐L1^High, CD11b⁻ CD103⁺ PD‐L1^Low and CD11b⁺ CD103⁻ PD‐L1^Int. Among them, the CD11b⁻ CD103⁺ PD‐L1^High DC subset highly induced Foxp3⁺ T cells. This subset expressed Aldh1a2 and Itgb8 genes, which are involved in retinoic acid metabolism and transforming growth factor‐β (TGF‐β) activation, respectively. Exogenous TGF‐β supplementation equalized the level of Foxp3⁺ T‐cell induction by the four subsets whereas retinoic acid did not, which suggests that high ability to activate TGF‐β is determinant for the high Foxp3⁺ T‐cell induction by CD11b⁻ CD103⁺ PD‐L1^High DC subset. Finally, this subset exhibited a migratory DC phenotype and could take up and present orally administered antigens. Collectively, the MLN CD11b⁻ CD103⁺ PD‐L1^High DC subset probably takes up luminal antigens in the intestine, migrates to MLNs, and highly induces regulatory T cells through TGF‐β activation.     

HnRNPM and CD44s expression affects tumor aggressiveness and predicts poor prognosis in breast cancer with axillary lymph node metastases

HnRNPM is an essential splicing factor and its expression is closely correlated with invasion and metastasis of tumor cells. The CD44 cell adhesion molecule is aberrantly expressed in many breast tumors and CD44 splice variants have been implicated in specific oncogenic signaling pathways. To investigate the clinical significance and biological function of hnRNPM, immunohistochemistry, quantitative, and semiquantitative polymerase chain reaction, lentiviral transfection system and transwell invasion assays were performed. We found that hnRNPM expression was significantly upregulated in breast cancer tissues compared with benign breast lesions. Although there was no significant correlation between hnRNPM and total CD44 protein or mRNA level, there was a negative correlation between hnRNPM and CD44v6. HnRNPM and CD44s expression showed positive correlation and in particular, they were dually expressed in breast cancer tissues. Interestingly, cancer stem cells marker, ALDH1⁺ phenotype was positively associated with overexpression of CD44s or hnRNPM and negatively related to CD44v6. Patients with high hnRNPM tended to have higher levels of CD44s, shorter overall survival (OS) and higher rates of lymph node metastases (LNM). Remarkably, Kaplan‐Meier and Cox regression analyses displayed that hnRNPM⁺ or CD44s^high was a poor prognostic factor for OS of patients with LNM. Upregulation of hnRNPM in MCF‐7 cells caused a significant increase in cell invasion, and this effect may occur through the regulation of CD44s expression. In conclusion, overexpression of hnRNPM promotes breast cancer aggressiveness by regulating the level of CD44s, indicates a poor prognosis for patients with LNM, and has potential as therapeutic targets.     

Inhibition of cancer stem cell-like properties and reduced chemoradioresistance of glioblastoma using microRNA145 with cationic polyurethane-short branch PEI

Glioblastomas (GBMs) are the most common primary brain tumors with poor prognosis. CD133 has been considered a putative marker of cancer stem cells (CSCs) in malignant cancers, including GBMs. MicroRNAs (miRNAs), highly conserved small RNA molecules, may target oncogenes and have potential as a therapeutic strategy against cancer. However, the role of miRNAs in GBM-associated CSCs remains mostly unclear. In this study, our miRNA/mRNA-microarray and RT-PCR analysis showed that the expression of miR145 (a tumor-suppressive miRNA) is inversely correlated with the levels of Oct4 and Sox2 in GBM-CD133⁺ cells and malignant glioma specimens. We demonstrated that miR145 negatively regulates GBM tumorigenesis by targeting Oct4 and Sox2 in GBM-CD133⁺. Using polyurethane-short branch polyethylenimine (PU-PEI) as a therapeutic-delivery vehicle, PU-PEI-mediated miR145 delivery to GBM-CD133⁺ significantly inhibited their tumorigenic and CSC-like abilities and facilitated their differentiation into CD133⁻-non-CSCs. Furthermore, PU-PEI-miR145-treated GBM-CD133⁺ effectively suppressed the expression of drug-resistance and anti-apoptotic genes and increased the sensitivity of the cells to radiation and temozolomide. Finally, the in vivo delivery of PU-PEI-miR145 alone significantly suppressed tumorigenesis with stemness, and synergistically improved the survival rate when used in combination with radiotherapy and temozolomide in orthotopic GBM-CD133⁺-transplanted immunocompromised mice. Therefore, PU-PEI-miR145 is a novel therapeutic approach for malignant brain tumors.     

Updated follow-up of a tolerance protocol in HLA-identical renal transplant pairs given donor hematopoietic stem cells

Kidney transplant recipients given donor hematopoietic stem cells from their HLA-identical living related donors have now been followed between 5 and 9½ years post-operatively. Recipients who were designated as tolerant (Tol) have remained so since the last report when the 5?year (biopsy associated) milestone was reached. There has been 1 mortality of a Tol patient, unrelated to the study protocol, while 5 (of 15) have remained Tol between 7 and 8½ years post-operatively. There has been continuing elevated T-regulatory (CD4⁺CD25^HighCD127^?FOXP3⁺) cells in PBMC previously reported on. Ten year renal transplant biopsies are tentatively planned.     

MEK-ERK signaling dictates DNA-repair gene MGMT expression and temozolomide resistance of stem-like glioblastoma cells via the MDM2-p53 axis

Overcoming the resistance of glioblastoma cells against temozolomide, the first-line chemotherapeutic agent of choice for newly diagnosed glioblastoma, is a major therapeutic challenge in the management of this deadly brain tumor. The gene encoding O(6) -methylguanine DNA methyltransferase (MGMT), which removes the methyl group attached by temozolomide, is often silenced by promoter methylation in glioblastoma but is nevertheless expressed in a significant fraction of cases and is therefore regarded as one of the most clinically relevant mechanisms of resistance against temozolomide. However, to date, signaling pathways regulating MGMT in MGMT-expressing glioblastoma cells have been poorly delineated. Here in this study, we provide lines of evidence that the mitogen-activated protein/extracellular signal-regulated kinase kinase (MEK)-extracellular signal-regulated kinase (ERK)-murine double minute 2 (MDM2)-p53 pathway plays a critical role in the regulation of MGMT expression, using stem-like glioblastoma cells directly derived from patient tumor samples and maintained in the absence of serum, which not only possess stem-like properties but are also known to phenocopy the characteristics of the original tumors from which they are derived. We show that, in stem-like glioblastoma cells, MEK inhibition reduced MDM2 expression and that inhibition of either MEK or MDM2 resulted in p53 activation accompanied by p53-dependent downregulation of MGMT expression. MEK inhibition rendered otherwise resistant stem-like glioblastoma cells sensitive to temozolomide, and combination of MEK inhibitor and temozolomide treatments effectively deprived stem-like glioblastoma cells of their tumorigenic potential. Our findings suggest that targeting of the MEK-ERK-MDM2-p53 pathway in combination with temozolomide could be a novel and promising therapeutic strategy in the treatment of glioblastoma.     

Toxic Epidermal Necrolysis in Polymedicated Patient Treated With Radiotherapy

Temozolomide is an oral alkylating agent indicated for the treatment of patients with glioblastoma multiforme concomitantly with radiotherapy and subsequently as monotherapy treatment. We report the case of a patient who developed toxic epidermal necrolysis (TEN) while she was being treated with chemoradiotherapy and several drugs. Cutaneous tests were performed with the drugs involved with negative result. Although the occurrence of TEN contraindicates suspected drug readministration, we based the decision to perform the controlled administration of temozolomide on the following reasons: (1) the poor prognosis of the underlying disease, (2) the lack of therapeutic alternatives, (3) the suspicion that other drugs taken by the patient simultaneously may be responsible (as anticonvulsants and trimethoprim sulfamethoxazole [TMP-SMX]), and (4) temozolomide was the first choice for treating the patient''s disease. The administration of a cumulative dose of 60 mg of temozolomide caused a slight skin reaction. Given this result, we conducted controlled administration of other drugs involved. Dexamethasone, codeine, omeprazole and levetiracetam were well tolerated. However, TMP-SMX produced a similar reaction to that caused by temozolomide. In conclusion, we present the first case of TEN induced by temozolomide and TMP-SMX associated with cranial radiotherapy confirmed by controlled administration. Radiotherapy in combination with these drugs could have favored TEN, as some authors have postulated, but we cannot prove this.     

Increased frequency of CD56Bright NK‐cells,CD3−CD16+CD56− NK‐cells and activated CD4+T‐cells or B‐cells in parallel with CD4+CDC25High T‐cells control potentially viremia in blood donors with HCV

A detailed phenotypic analysis of major and minor circulating lymphocyte subsets is described in potential blood donors with markers of hepatitis C virus (HCV), including non‐viremic and viremic groups. Although there were no changes in the hematological profile of either group, increased the levels of pre‐NK cells (CD3^?CD16⁺CD56^?) and a lower frequency of mature NK cells (CD3^?CD16⁺CD56⁺) characterized innate immunity in the non‐viremic group. Both non‐viremic and viremic groups displayed significantly increased levels of CD56^Bright NK cells. Furthermore, this subset was significantly elevated in the viremic subgroup with a low viral load. In addition, an increase in the NKT2 subset was observed only in this subgroup. An enhanced frequency of activated CD4⁺ T‐cells (CD4⁺HLA‐DR⁺) was a characteristic feature of the non‐viremic group, whereas elevated CD19⁺ B‐cells and CD19⁺CD86⁺ cell populations were the major phenotypic features of the viremic group, particularly in individuals with a low viral load. Although CD4⁺CD25^High T‐cells were significantly elevated in both the viremic and non‐viremic groups, it was particularly evident in the viremic low viral load subgroup. A parallel increase in CD4⁺CD25^High T‐cells, pre‐NK, and activated CD4⁺ T‐cells was observed in the non‐viremic group, whereas a parallel increase in CD4⁺CD25^High T‐cells and CD19⁺ B‐cells was characteristic of the low viral load subgroup. These findings suggest that CD56^Bright NK cells, together with pre‐NK cells and activated CD4⁺ T‐cells in combination with CD4⁺CD25^High T‐cells, might play an important role in controlling viremia. Elevated CD56^Bright NK cells, B‐cell responses and a T‐regulated immunological profile appeared to be associated with a low viral load. J. Med. Virol. 81:49–59, 2009. © 2008 Wiley‐Liss, Inc.     

The origin of regulatory from the effector cells in LAG-3-marked Th1 immunity against severe influenza virus infection

In severe respiratory virus infections, including influenza, an exaggerated host immune response has been linked to the severe disease and death. Control of the overwhelming immune response is thus essential. Efforts with broad-spectrum immunosuppressive agents such as steroids are disappointing. A better understanding of host immune response using animal experimental system is required to avoid undesired outcome of experimental manipulation. Following severe influenza virus infection in influenza hemagglutinin antigen-specific transgenic mouse experimental model, step-wise evolving cells from a pool of naïve hemagglutinin-specific CD4⁺ T cells were studied for phenotypic, genomic, and functional characterization in vivo. Naïve CD4⁺ T cells respond with Th1 commitment in the absolute majority. They first develop into LAG-3^MedIFN-γ-secreting Th1 effectors and then evolve into LAG-3^HighIFN-γ-not-secreting regulators with increasing LAG-3 expression upon continuous activation and cell division. The LAG-3^MedIFN-γ-secreting effectors contribute to inflammation, boost inflammatory response of cognate antigen-specific CD8⁺ T cells, and aggravate the disease despite facilitated virus clearance. In contrast, LAG-3^High regulators do not contribute to inflammation, suppress CD8⁺ T cell inflammatory response, alleviate lung pathology, and ameliorate the disease with preserved virus clearance. Moderated CD8⁺ T cells retain proliferative capacity, and persist beyond virus clearance. Such moderation is distinct from Foxp-3⁺ regulator-mediated suppression, which suppresses proliferative and inflammatory responses of the CD8⁺ T cells and impairs virus clearance with inflammation alleviation. Origin of regulatory from the effector cells of LAG-3-marked Th1 immunity alleviates lung inflammation without impairment of virus eradication.     

The role of ALDH1A1 in contributing to breast tumour aggressiveness: A study conducted in an African population

Aldehyde dehydrogenase 1 member A1 (ALDH1A1) is one of the most well studied breast cancer stem cells. Its expression has been associated with poor clinicopathological features and clinical outcomes in several studies. This paper studies the expression of ALDH1A1 and its combination with CD44⁺/CD24^−/low breast cancer stem cell and their association with clinicopathological parameters and molecular subtypes.MethodTissue Microarray was constructed from 222 Formalin Fixed Paraffin Embedded (FFPE) breast cancer tissues. The expression of ALDH1A1, CD44 and CD24 were assessed by Immunohistochemistry (IHC). The association of ALDH1A1 and its association with clinicopathological parameters, molecular subtypes, CD44 and CD24 were studied in an African population. The association between CD44⁺/CD24^−/low/ALDH1⁺ and the clinicopathological phenotypes were also studied.ResultsA high ALDH1A1 expression of 90% was recorded in this study. No association was found between ALDH1A1 and clinicopathological parameters. ALDH1A1 was positively associated with CD24 (r = 0.228, OR-4.599 95% CI- 1.751–12.076, p = 0.001) and CD44 (r = 0.228, OR-5.538 95%CI- 1.841–16.662, p = 0.001) but not associated with CD44⁺/CD24^−/low (r = 0.134, OR- 2.720 95%CI- 0.959–7.710, p = 0.052). CD44⁺/CD24⁻/ALDH1⁺ however had significant associations with Age (p- 0.020, r = 0.161, OR- 2.771, 95%CI 1.147–6.697), Gender (p = 0.004, OR- 15.333 95%CI 1.339–175.54), Tumour grade (p = 0.005, r = 0.197, OR-3.913 95%CI 1.421–10.776) and clinical prognostic staging (p = 0.014, r = 0.182, OR-3.028 95%CI- 1.217–7.536). There was no association between CD44⁺/CD24⁻/ALDH1⁺ and the molecular subtypes.ConclusionThe high expression of ALDH1A1 in breast cancer makes it an important target for targeted therapy. This study further confirms the increased tumourigenicity of CD44⁺/CD24⁻/ALDH1⁺ combination phenotype and its association with increased tumour grade and clinical prognostic stage. Survival studies of ALDH1A1 and other breast cancer stem cells in African populations are strongly recommended to help further understand their effect on tumour aggressiveness.     

肿瘤相关巨噬细胞在晚期卵巢癌组织中的浸润与肿瘤浸润淋巴细胞表型、免疫效能之间的关系

目的:探讨晚期卵巢癌组织中浸润的肿瘤相关巨噬细胞(tumor-associated macrophage,TAM)与肿瘤浸润淋巴细胞(tumor-infiltrating lymphocyte,TIL)表型及免疫效能的关系。方法:免疫组化方法分析175例低分化卵巢癌组织病理切片中TAM分布密度,以中位数为界限将病例分为TAM高密度组和TAM低密度组,对照组为32例良性卵巢病变组织;应用流式细胞术分析TAM高密度组与TAM低密度组中TIL的CD8⁺和CD25⁺表型变化情况;体外扩增培养TIL后取细胞培养上清液,ELISA法分析各组TIL中白细胞介素2(IL-2)、白细胞介素-10(IL-10)、转化生长因子β(TGF-β)和干扰素γ(IFN-γ)细胞因子表达变化。结果:175例低分化卵巢癌组织中TAM平均浸润密度为62.8/高倍镜视野(HP,×400),中位数为53.3/HP,其中TAM高密度组87例,TAM低密度组88例;对照组TAM平均浸润密度10.5/HP(P<0.05)。CD8⁺在TAM高密度组中表达平均值为24%,在TAM低密度组中表达平均值为52%(P<0.05);CD25⁺在TAM高密度组中表达平均值为48%,在TAM低密度组中表达平均值为25%(P<0.05);对照组中CD8⁺和CD25⁺的TIL平均浸润密度为7%, TAM高密度组及TAM低密度组中CD8⁺和CD25⁺的TIL平均浸润密度显著高于对照组(P<0.05)。与TAM低密度组比较,TAM高密度组中TIL的杀伤性细胞因子IL-2和IFN-γ表达明显减少(P<0.05),而抑制性细胞因子IL-10和TGF-β表达明显增加(P<0.05)。结论:高密度TAM浸润的卵巢癌组织中,CD25⁺的TIL表型增多,CD8⁺的TIL表型减少,抑制性细胞因子IL-10和TGF-β表达增加,杀伤性细胞因子IL-2和IFN-γ表达减少,提示TAM浸润密度与TIL表型及免疫效能相关。     

Fatty acid binding protein 7 as a marker of glioma stem cells

Glioblastomas are the most aggressive brain tumors. Glioblastoma stem cells (GSCs) are thought to be responsible for the recurrence, chemoresistance, and poor prognosis of glioblastoma. Fatty acid binding protein 7 (FABP7), which is a cellular chaperone for a variety of omega‐3 fatty acids, is a known marker for neural stem cells. In this study, using a newly developed anti‐FABP7 antibody and patient‐derived GSC lines, we evaluated the expression of FABP7 in GSCs. Using immunocytochemistry, Western blotting, and qPCR analyses, FABP7 was found to be highly enriched in GSCs and its localization was found in cytosol and nuclei. FABP7 expression was significantly downregulated in differentiated GSCs induced by the addition of serum. In the glioma surgical specimens, FABP7 was highly expressed in the majority of glioblastoma. Double immunostaining for FABP7 and Sox2 showed that FABP7⁺Sox2⁺ tumor cells were significantly increased in glioblastoma (grade IV) compared with diffuse astrocytoma (grade II) and anaplastic astrocytoma (grade III). Our data introduces FABP7 as a marker for GSCs and further highlights its possible significance for glioma diagnosis and treatment.     

Cooperative action of CD8 T lymphocytes and natural killer cells controls tumour growth under conditions of restricted T‐cell receptor diversity

In mice expressing a transgenic T-cell receptor (TCR; TCRP1A) of DBA/2 origin with reactivity towards a cancer-germline antigen P1A, the number of TCRP1A CD8⁺ T cells in lymphoid organs is lower in DBA/2 than in B10.D2 or B10.D2(× DBA/2)F₁ mice. This reduction results from haemopoietic cell autonomous differences in the differentiation of the major histocompatibility complex class I-restricted TCRP1A thymocytes controlled by DBA/2 versus B10.D2-encoded genes. We report here that the lower number of TCRP1A CD8⁺ T cells in DBA/2 mice correlated with their poor resistance to P1A-expressing mastocytoma solid tumours. Functional potency of CD8⁺ cytolytic T lymphocytes (CTL) from the above strains was not compromised, but their number after expansion appeared to be influenced by their genetic background. Intriguingly, non-transgenic DBA/2 mice resisted P1A⁺ tumours more efficiently despite poor representation of P1A-specific CTL. This was partly the result of their more heterogeneous TCR repertoire, including reactivity to non-P1A tumour antigens because mice that had rejected a P1A⁺ tumour became resistant to a P1A⁻ variant of the tumour. Such ‘cross-resistance’ did not develop in the TCRP1A transgenic mice. Nonetheless, reconstitution of RAGº^/º mice with TCRP1A CD8⁺ T cells, with or without CD4⁺ T cells, or exclusive representation of TCRP1A CD8⁺ T cells in RAGº^/º TCRP1A transgenic mice efficiently resisted the growth of P1A-expressing tumours. Natural killer cells present at a higher number in RAGº^/º mice also contributed to tumour resistance, in part through an NKG2D-dependent mechanism. Hence, in the absence of a polyclonal T-cell repertoire, precursor frequencies of natural killer cells and tumour-specific CTL affect tumour resistance.