补骨脂与双氢青蒿素合剂治疗小鼠隐孢子虫病的实验研究

用补骨脂和双氢青蒿素合剂治疗经地塞米松诱导的隐孢子虫病小鼠, 剂量为补骨脂3.4 g/kg, 双氢青蒿素60 mg/(kg·d), 灌胃, 连用7 d。 结果粪检隐孢子虫卵囊数、 全血CD4⁺、 CD3⁺ T细胞比例、 血清γ干扰素 (IFN-γ) 浓度, 以及小肠组织匀浆一氧化氮 (NO) 浓度均高于感染对照组 (P值均＜0.01)。小肠组织病变程度均轻于感染对照组。表明补骨脂和双氢青蒿素合剂可通过调节血清IFN-γ、 全血CD4⁺、 CD3⁺ T 细胞比例, 以及提高肠组织NO浓度, 参与宿主免疫应答和炎症反应过程, 抑杀虫体、 治疗小鼠隐孢子虫病。     

Prevention of cyclophosphamide-induced accelerated diabetes in the NOD mouse by nicotinamide or a soy protein-based infant formula

Spontaneous diabetes in the NOD mouse can be prevented by nicotinamide or by an infant formula diet in which the protein source is replaced with casein hydrolysate (Pregestimil) or soy protein (Prosobee). NOD mice maintained on the standard diet (chow and water) and given cyclophosphamide (Cy) at day 95 develop accelerated and synchronised diabetes within 14 days. Here, we compared the ability of oral nicotinamide or Prosobee, either given alone or concurrently, from weaning, in preventing diabetes in the Cy model. The resulting insulitis and the expression of intra-islet inducible nitric oxide synthase (iNOS) were examined at days 0, 4, 7, 11 and 14 following Cy administration. Intra-islet CD4 and CD8 cells and macrophages were also enumerated at day 11. In mice given the standard diet and injected with Cy at day 95 (group 5), diabetes developed in 7/11 mice, 14 days later. Mice exposed to oral nicotinamide (group 2), Prosobee (group 3) or both (group 4), did not develop the disease during this period and until a further 30 days (p = 0.03). In mice exposed to the standard diet and without Cy treatment (group 1) the insulitis scores increased slowly until day 11 and then declined slightly at day 14 whereas mice exposed to the same diet but given Cy at day 95, showed a sharp decline at day 4 followed by a rapid increase between day 7-14. However, in mice given either nicotinamide, Prosobee or both, the insulitis scores at most time-points were generally lower than in Cy-treated animals on the standard diet. In the latter group, CD4 and CD8 cells and macrophages were also higher at day 11 than all other 4 groups (CD4: p < 0.05; CD8: p< 0.05; macrophages: p<0.0001). The number of iNOS labelled cells increased progressively in mice on the standard diet and given Cy and were significantly higher at days 4, 7 and 11 than in the 3 dietary groups. Thus, oral nicotinamide or Prosobee, either alone or together, prevents Cy induced diabetes in the NOD mouse. The protective diets suppress Cy-induced intra-islet immune cell influx and iNOS expression.     

CD25+ cell depletion hastens the onset of severe disease in collagen-induced arthritis

OBJECTIVE: CD4+,CD25+ T regulatory cells may offer opportunities to intervene in the course of autoimmune disease. We wished to evaluate their potential for influencing systemic and chronic joint inflammation by investigating their involvement in collagen-induced arthritis (CIA). METHODS: We depleted DBA/1 mice of CD25+ regulatory cells by injection of a depleting monoclonal antibody specific for CD25 14 days before a single immunization with type II collagen (CII) in Freund's complete adjuvant. CD4+,CD25+ T cells were adoptively transferred to some groups of mice during immunization. Mice were then scored for signs of arthritis, and blood was taken periodically to measure the amounts of CII-specific antibodies. Splenocytes of treated mice were examined in vitro to determine the effects of depletion on proliferation to CII and control antigens. RESULTS: CD25+ cell-depleted DBA/1 mice had significantly more severe disease than control mice following collagen immunization. The magnified severity was also accompanied by higher antibody titers against collagen, and in vitro tests showed increased proliferation of collagen-specific T cells. Adoptively transferring CD4+,CD25+ T cells into depleted mice was shown to reverse the heightened severity. Control mice, which were depleted and immunized with the neoantigen keyhole limpet hemocyanin (KLH), had neither an increased antibody response toward KLH nor an augmented proliferative response, indicating that CD25+ cell depletion preferentially affects immunity against self antigen. CONCLUSION: These results establish a link between CD4+,CD25+ regulatory cells and CIA and provide a rationale for investigating CD4+,CD25+ T regulatory cells in the treatment and prevention of arthritis.     

Decreased haem oxygenase-1 and increased inducible nitric oxide synthase in the lung of severe COPD patients.

Oxidant/antioxidant imbalance is implicated in the pathogenesis of chronic obstructive pulmonary disease (COPD). The current study examined the expression of antioxidant and pro-oxidant enzymes, haem oxygenases (HO) and inducible nitric oxide synthase (iNOS) respectively, in patients with severe COPD and control smokers without lung function impairment. Immunoreactivity for HO-1, HO-2, iNOS and nitric oxide-derived oxidants expressed as nitrotyrosine (N-Tyr) was quantified in peripheral lung. HO-1+ alveolar macrophages were decreased in severe COPD compared to control smokers, whereas no difference was observed in iNOS+ macrophages. In contrast, severe patients had significantly higher numbers of iNOS+ cells in alveolar walls. These iNOS+ cells were identified as type 2 pneumocytes and their number was inversely related to HO-1+ macrophages. There were no significant differences in N-Tyr immunostaining between the two groups. However, the rate of protein nitration in lung tissue was directly related to iNOS expression and associated with lower values of forced expiratory volume in one second/forced vital capacity. HO-2 was constitutively expressed by type 2 pneumocytes and these cells were increased in severe COPD. In conclusion, the results suggest that the enzymes involved in the oxidative stress response may have a different role in the lung defence and that imbalance between haem oxygenase-1 and inducible nitric oxide synthase may be associated with the development of severe impairment in chronic obstructive pulmonary disease patients.     

Changes in the expression and distribution of the inducible and endothelial nitric oxide synthase in mucosal biopsy specimens of inflammatory bowel disease

OBJECTIVE: The role of nitric oxide (NO) in the pathophysiology of inflammatory bowel disease (IBD) is controversial. The aim of this study was to investigate the expression and localization of nitric oxide synthase isoforms (iNOS, eNOS) in IBD colonic mucosa. MATERIAL AND METHODS: Forty-four patients with IBD (24 ulcerative colitis (UC), 20 Crohn's disease (CD) and 16 controls) were investigated by colonoscopy. iNOS and eNOS in tissue sections was demonstrated by histochemistry (NADPH-diaphorase reaction) and immunohistochemistry. Cell type analysis and quantitative assessment of the iNOS immunoreactive (IR) cells and densitometry of iNOS in immunoblots were also performed. RESULTS: iNOS-IR cells were significantly numerous in inflamed mucosa of UC (64+/-4 cells/mm2) than in CD (4+/-2 cells/mm2). iNOS-IR/CD15-IR cells showed significant elevation in inflamed (i) versus uninflamed (u) UC mucosa (UCu 8+/-3%, UCi 85+/-10%) In CD, the percentage of iNOS-IR/CD68-IR cells was lower in inflamed sites (CDu 23+/-8%, CDi 4+/-3%). Immunoblot of biopsies revealed significant elevation of iNOS in active UC compared with uninflamed sites, whereas in CD no significant changes were detected. Differences were observed in eNOS and endothelial marker CD31 immunoreactivity. In patients with UC and in controls the ratios of eNOS/CD31-IR vessels were 82.3% and 92.0% respectively, whereas in CD the ratio was 8.3% with a concomitantly significant increase of CD31-IR vessels. The distribution and morphological characteristics of the NOS-IR inflammatory cells and endothelia were similar to those showing NADPH-diaphorase reactivity. CONCLUSIONS: Differences observed in the expression and distribution of NOS isoforms in immune and endothelial cells may contribute to better understanding of the structural and physiological changes in UC and CD.     

TACE联合自体CIK治疗原发性肝癌患者的细胞免疫功能变化

目的 探讨肝动脉灌注化学治疗栓塞(TACE)联合细胞因子诱导的杀伤细胞(CIK)治疗原发性肝癌(PHC)患者的细胞免疫功能变化.方法 65例PHC患者分为2组.对照组和研究组分别为32例和33例.单纯TACE治疗组(对照组)常规行肝动脉灌注化学治疗栓塞术;TACE联合细胞免疫治疗组(研究组)采集外周血单个核细胞(PBM...     

TNF-α, nitric oxide and IFN-γ are all critical for development of necrosis in the small intestine and early mortality in genetically susceptible mice infected perorally with Toxoplasma gondii

We previously reported that genetic susceptibility of mice to peroral infection with T. gondii is associated with CD4+ T cell-dependent, interferon (IFN)-gamma-mediated necrosis of their small intestine. We examined the role of tumour necrosis factor (TNF)-alpha and nitric oxide (NO), in addition to IFN-gamma. At 7 days after infection, a marked increase in CD4+ T cells was observed in lamina propria mononuclear cells (LPC) of the small intestine as compared with normal mice, and significantly greater amounts of mRNA for IFN-gamma, TNF-alpha, and inducible NO synthase (iNOS) were detected in LPC of the small intestine of infected than uninfected animals. Treatment of infected mice with anti-TNF-alpha monoclonal antibody (mAb) or the iNOS inhibitor, aminoguanidine, prevented necrosis and prolonged time to death. Infected iNOS-targeted mutant mice did not develop the disease whereas infected, control mice did. Treatment with anti-TNF-alpha mAb did not affect the expression of IFN-gamma in the LPC but inhibited expression of iNOS in the infected mice, indicating the role of TNF-alpha in the induction of iNOS. These results suggest that NO induced by a combination of IFN-gamma and TNF-alpha through activation of iNOS is a critical mediator of intestinal pathology and contributes to early mortality in genetically susceptible mice.     

Effect of IL-18 on leukocyte expression of iNOS and phospho-IkB in patients with squamous cell carcinoma of the oral cavity

The inducible synthase of nitric oxide (iNOS) is responsible for the synthesis of nitric oxide (NO) in neutrophils (PMN) and in peripheral blood mononuclear cells (PBMC). This enzyme is controlled by a number of cytokines which accomplish their biological effect via e.g. activation of NF-kB pathway. The aim of the present study was to assess the expression of iNOS and production of NO by PMN and PBMC in patients with oral cavity squamous cell carcinoma (SCC), and to examine the role of the NF-kB pathway in the IL-18-stimulated activation of this enzyme. The production of NO and iNOS expression in PMN were reduced, while iNOS expression in PBMC was increased but NO production by these cells remain unchanged. Patients after treatment showed lower intensity of iNOS expression compared to that observed before treatment. Moreover, both before and after treatment iNOS expression was inversely correlated with phospho-IkB expression in PMN and in PBMC. Significantly higher levels of total NO were observed in the serum of Stage IV patients before and after treatment as compared to the control group. Altered expression of iNOS and NO generation by PMN and PBMC may have an unfavorable effect on the course of antineoplastic response in patients with squamous cell carcinoma of the oral cavity. Intensification of iNOS expression and NO production in Stage IV patients, induced by rhIL-18, suggests its beneficial effect on the activity of leukocytes in patients with squamous cell carcinoma of the oral cavity.     

Estrogen up-regulates inducible nitric oxide synthase, nitric oxide, and cyclooxygenase-2 in splenocytes activated with T cell stimulants: role of interferon-gamma

Estrogen is implicated in many autoimmune diseases and is a robust immunomodulator. For example, it regulates interferon (IFN)-gamma, a cytokine believed to up-regulate inducible nitric oxide synthase (iNOS). A notable gap in the literature is a lack of information on the regulation of nitric oxide in immune tissues by estrogen. We now show that activation of splenocytes with T cell stimulants [concanavalin-A (Con-A) or anti-CD3 antibodies] results in copious release of nitric oxide in splenocyte cultures from estrogen-treated but not placebo-treated mice. Moreover, even a low dose of T cell stimulants induced nitric oxide in splenocytes from estrogen-treated, but not placebo-treated, mice. Con-A-activated splenocytes from estrogen-treated mice also have up-regulated iNOS mRNA, iNOS protein, and cyclooxygenase-2 (a nitric oxide-regulated downstream proinflammatory protein) when compared with controls. Our studies suggest that the induction of nitric oxide by activated splenocytes from estrogen-treated mice is mediated in part by IFNgamma. First, blocking costimulatory signals mediated through interactions of CD28 and B7 molecules by CTLA-4Ig markedly decreased not only IFNgamma but also nitric oxide. Second, estrogen treatment of IFNgamma-knockout (IFNgamma(-)/(-)) mice did not induce iNOS protein or nitric oxide. Finally, in vitro addition of recombinant IFNgamma to Con-A-activated splenocytes from IFNgamma((-)/(-)) mice induced iNOS protein primarily in estrogen-treated mice. Overall, this is the first report to show that estrogen treatment up-regulates IFNgamma-inducible-iNOS gene expression, iNOS protein, nitric oxide, and cyclooxygenase-2 as an indirect consequence of activation of T cells. These findings may have wide implications to immunity and inflammatory disorders including female-predominant autoimmune diseases.     

Immune reconstitution following allogeneic peripheral blood progenitor cell transplantation: comparison of recipients of positive CD34+ selected grafts with recipients of unmanipulated grafts

We compared the kinetic recovery of lymphocytes and their subsets in two groups of patients submitted to allogeneic peripheral blood progenitor cell transplantation (allo-PBT): those receiving lymphocyte-depleted leukaphereses by positive selection of CD34+ cells (group 1, n = 18) and those receiving unmanipulated leukaphereses (group 2, n = 15). Patients were conditioned with cyclophosphamide (120 mg/kg) and fractioned total body irradiation (13 Gy, group 1; 12 Gy, group 2). The mean number (x 10(6)/kg) of CD34+ and CD3+ cells infused was 4.0 and 0.67, respectively, in group 1 patients, and 4.7 and 274, respectively, for group 2 patients. Graft-versus-host disease prophylaxis consisted of cyclosporin A + methylprednisolone for group 1 and cyclosporin A + methotrexate for group 2. Median follow-up was 7 months (range 2-8 months) for both groups. During the first 6 months post-transplant, CD4+ cell counts were lower in group 1 as compared with group 2 (p = 0.014, 0.010, 0.011, 0.0003, and 0.052 at 0.5, 1, 2, 3, and 6 months, respectively), whereas there was no difference at 8 months. The number of CD4+CD45RA+ cells was very low throughout the study in both groups, being lower in group 1 than in group 2, especially during the first 3 months post-transplant (p = 0.007 and 0.0006 at 1 and 3 months). Normal levels of CD8+ cells were reached by 1 month post-transplant in both groups. TCR gamma delta + cell counts were lower in group 1 than in group 2 during the first 4 months post-transplant (p = 0.001, 0.004, and 0.04 at 1, 3, and 4 months). A normal number of natural killer cells (CD3-CD56+) was achieved 1 month post-transplant in both groups. B lymphocytes (CD19+) showed low or undetectable counts throughout the first 4 months in both groups, achieving the normal range at 8 months. These results show that, during the first 6 months following allo-PBT with CD34+ selected grafts, the number of CD4+, CD4+CD45RA+, and TCR gamma delta + cells is significantly lower than after unmanipulated allo-PBT; these differences disappeared at 8 months. In contrast, there are no differences between transplant groups in the recovery of CD8+, CD19+, and natural killer cells.     

Thoracoscopic talc poudrage decreases T-lymphocytes in the peripheral blood

BACKGROUND: Thoracoscopic talc poudrage induces peripheral blood granulocytosis and lymphopenia. The aim of this study is to investigate the type of lymphopenia in patients undergoing thoracoscopic talc poudrage. METHODS: We have measured peripheral blood lymphocyte subsets in 11 patients undergoing thoracoscopic talc poudrage, before (baseline), at 24 and 48 h after the procedure. Lymphocyte numbers were analysed by flow cytometry for the evaluation of the CD3+, CD4+, CD8+ cells (total T-lymphocytes, helper T-lymphocytes, cytotoxic T-lymphocytes, respectively), the CD19+ cells (B-lymphocytes), and the CD16+, CD56+ and CD57+ cells (NK-cells). No anti-inflammatory medication was permitted before, during or after the procedure. RESULTS: Absolute peripheral blood lymphocyte count significantly decreased following thoracoscopic talc poudrage compared to baseline values (p=0.007). Similarly, peripheral blood CD3+, CD4+ and CD8+ lymphocyte counts significantly decreased compared to baseline (p=0.005, 0.02 and 0.03, respectively) with a more prominent reduction of CD3/CD45RO memory cells. No significant difference was found in the absolute number of CD19+, CD16+, CD56+, and CD57+ cells before and after thoracoscopic talc poudrage. CONCLUSION: Patients undergoing thoracoscopic talc poudrage display peripheral blood T-lymphopenia following the procedure.     

The pattern of immune cell infiltration in chromoblastomycosis: involvement of macrophage inflammatory protein-1 alpha/CCL3 and fungi persistence

Chromoblastomycosis (CR) is a subcutaneous chronic mycosis characterized by a granulomatous inflammatory response. However, little is known regarding the pattern of leukocyte subsets in CR and the pathways involved in their recruitment. The objective of this study was to assess the cellular subsets, chemokine, chemokine receptors and enzymes in CR. The inflammatory infiltrate was characterized by immunohistochemistry using antibodies against macrophages (CD68), Langerhans'cells (S100), lymphocytes (CD3, CD4, CD8, CD45RO, CD20 and CD56) and neutrophils (CD15). The expression of MIP-1alpha (Macrophage inflammatory protein-1alpha), chemokine receptors (CXCR3 and CCR1) and enzymes (superoxide dismutase-SOD and nitric oxide synthase-iNOS) was also evaluated by the same method. We observed an increase in all populations evaluated when compared with the controls. Numbers of CD15(+) and CD56(+) were significantly lower than CD3(+), CD4(+), CD20(+) and CD68(+) cells. Statistical analysis revealed an association of fungi numbers with CD3, CD45RO and iNOS-positive cells. Furthermore, MIP-1alpha expression was associated with CD45RO, CD68, iNOS and CXCR3. Our results suggest a possible role of MIP-1alpha and fungi persistence in the cell infiltration in CR sites.     

Dual colour fluorescein analysis of peripheral blood T cells in auto-immune chronic active hepatitis

Both CD4 and CD8 T cells are subdivided into two phenotypically distinct sublineages via another two T cell markers, Leu-8 and CD11b antigens. The proportions of these four T cell subsets, CD4+Leu8+, CD4+Leu8-, CD8+11b+ and CD8+11b-, were studied in patients with auto-immune chronic active hepatitis (CAH) and compared with disease controls (hepatitis B surface antigen positive chronic active hepatitis) and healthy controls. We found that the proportion of CD4+Leu8+ cells was significantly reduced compared with controls (P less than 0.01), whereas those of the other cells were almost identical in all 3 groups. The absolute number of these CD4+Leu8+ cells was also lower than that of controls (P less than 0.01). Thus, the present study suggests that a reduced number of CD4+Leu8+ cells is associated with the aberrant immune response in auto-immune CAH.     

Changes in the expression and distribution of the inducible and endothelial nitric oxide synthase in mucosal biopsy specimens of inflammatory bowel disease

Objective The role of nitric oxide (NO) in the pathophysiology of inflammatory bowel disease (IBD) is controversial. The aim of this study was to investigate the expression and localization of nitric oxide synthase isoforms (iNOS, eNOS) in IBD colonic mucosa. Material and methods Forty-four patients with IBD (24 ulcerative colitis (UC), 20 Crohn's disease (CD) and 16 controls) were investigated by colonoscopy. iNOS and eNOS in tissue sections was demonstrated by histochemistry (NADPH-diaphorase reaction) and immunohistochemistry. Cell type analysis and quantitative assessment of the iNOS immunoreactive (IR) cells and densitometry of iNOS in immunoblots were also performed. Results iNOS-IR cells were significantly numerous in inflamed mucosa of UC (64±4 cells/mm²) than in CD (4±2 cells/mm²). iNOS-IR/CD15-IR cells showed significant elevation in inflamed (i) versus uninflamed (u) UC mucosa (UC_u 8±3%, UC_i 85±10%) In CD, the percentage of iNOS-IR/CD68-IR cells was lower in inflamed sites (CD_u 23±8%, CD_i 4±3%). Immunoblot of biopsies revealed significant elevation of iNOS in active UC compared with uninflamed sites, whereas in CD no significant changes were detected. Differences were observed in eNOS and endothelial marker CD31 immunoreactivity. In patients with UC and in controls the ratios of eNOS/CD31-IR vessels were 82.3% and 92.0% respectively, whereas in CD the ratio was 8.3% with a concomitantly significant increase of CD31-IR vessels. The distribution and morphological characteristics of the NOS-IR inflammatory cells and endothelia were similar to those showing NADPH-diaphorase reactivity. Conclusions Differences observed in the expression and distribution of NOS isoforms in immune and endothelial cells may contribute to better understanding of the structural and physiological changes in UC and CD.     

不同阶段慢性乙型肝炎患者外周血单个核细胞中NF-κB/iNOS的表达及免疫状态

目的研究核因子-κB(NF-κB)/诱导型一氧化氮合成酶(iNOS)在不同阶段慢性乙型肝炎患者外周血单个核细胞(PBMC)中的表达,比较各阶段自然杀伤细胞及T、B淋巴细胞的变化情况。方法纳入福建医科大学附属南平第一医院2017年10月-2019年1月慢性乙型肝炎相关疾病患者,按照疾病不同阶段分4组:肝炎组、肝衰竭组、肝硬化组、肝癌组,每组各20例;另选取10例健康志愿者作为对照组。采用流式细胞仪检测5组外周血总淋巴细胞计数和淋巴细胞亚群计数;采用Western Blot检测PBMC中NF-κB、iNOS蛋白的表达水平;硝酸还原酶法和ELISA法检测血清中NO、IL-6水平。多组间比较采用单因素方差分析,进一步两两比较采用LSD-t检验。结果与对照组比较,肝炎组、肝衰竭组、肝癌组、肝硬化组总淋巴细胞计数水平均明显下降(P值均<0.05);与肝炎组比较,肝衰竭组、肝癌组、肝硬化组总淋巴细胞计数水平均明显下降(P值均<0.05)。肝衰竭组、肝癌组、肝硬化组CD3+明显下降,分别与对照组、肝炎组比较差异均有统计学意义(P值均<0.05);肝衰竭组、肝癌组、肝硬化组与肝炎组CD4+比较明显下降(P<0.05);肝衰竭组与肝癌组、肝硬化组、对照组CD8+比较明显升高(P值均<0.05);肝衰竭组较对照组、肝炎组、肝硬化组、肝癌组CD4+/CD8+明显降低(P值均<0.05);肝衰竭组、肝硬化组CD3-CD19+明显升高,分别与肝炎组、对照组比较差异均有统计学意义(P值均<0.05);肝衰竭组、肝癌组CD16+CD56+明显下降,分别与肝炎组、对照组比较差异均有统计学意义(P值均<0.05)。肝硬化组、肝衰竭组、肝癌组NF-κB、iNOS蛋白表达水平均明显增高,分别与肝炎组、对照组比较差异均有统计学意义(P值均<0.05)。与对照组比较,肝衰竭组、肝癌组、肝硬化组和肝炎组的NO和IL-6水平均明显升高(P值均<0.05)。结论不同阶段HBV感染者免疫状态存在一定差异性,NF-κB-iNOS-NO可能参与HBV相关疾病的免疫反应。     

Different patterns of some systemic immunological cell markers in HIV only, and HIV/hepatitis C-infected children

In three groups of children, aged 4-6 years (i.e., human immunodeficiency virus [HIV]-negative controls, HIV-seropositive, and dually HIV/hepatitis C virus (HCV)-seropositive), two types of immunological investigations in blood cells were performed: (a) numerical assays, consisting of flow cytometric measurement of different lymphocyte sets or subsets, as follows: CD3+, CD19+, CD4+, CD16+/CD56+; (b) functional assays, consisting of interleukin-1 (IL-1) levels as well as natural killer (NK)-cell dependent cytotoxicity, in CD14+, or CD16+/CD56+ sorted cells, respectively. Results revealed, in addition to the classic markers (i.e., lower numbers of CD4+ cells and a decreased CD4+/CD8+ ratio in both infected groups of subjects) other findings, as follows: increased numbers of CD8+ cells in dually infected children, accompanied by a lower CD4+/CD8+ ratio, as compared to HIV-infected alone; diminished numbers of CD16+/CD56+ cells in both groups of infected patients were correlated with a lower NK-cell cytotoxicity rate; a reduced capacity for IL-1 synthesis of sorted macrophages both in HIV-only and in HIV/HCV-seropositive subjects, but significantly more marked in dually infected children. The importance of the present data in the immune monitoring of AIDS disease in a pediatric population is discussed.     

Analysis of bone marrow and peripheral blood immunoregulatory lymphocytes in patients with myelodysplastic syndrome

The cell surface phenotype of immunoregulatory lymphocytes in bone marrow (BM) and peripheral blood (PB) in myelodysplastic syndrome (MDS), a stem cell disorder, was analyzed. Mononuclear cells from 25 patients with refractory anemia (RA) and nine with RA with an excess of blasts (RAEB) were characterized by two-color flow cytometry using various monoclonal antibodies. No significant change of CD3+, CD4+, and CD8+ cells in PB, but a decrease of the percent of positive cells for CD8+ among the total lymphocyte (%CD8+ +) was noticed in RA patients. On the other hand, in BM of RA patients, a decrease in the number of CD4+cells, but not CD8+ +cells, was noted. In RAEB patients, the absolute numbers of CD3+, CD4+, CD8+, and CD8+ +cells in BM were decreased; however, the ratio of these lymphocytes was not changed. No change was observed among the CD4 + subsets in PB of RA or RAEB patients. In BM, a decrease in percentage of CD4+ CD45RA+ (% CD4+ CD45RA+; naive cell) and increases in CD4+ CD45RO+ (% CD4+ CD45RO+; memory cell) and CD4+ CD29+ (%CD4+ CD29+; helper/inducer) among CD4+ cells were found in both RA and RAEB patients. Analysis of the CD8+ + subset showed an increased number of CD8+ + CD11a+ cells (activated CTL) in both BM and PB of RA patients, but not of RAEB patients. Furthermore, increments in CD56+ and CD16+ cells among CD3- cells (natural killer; NK cells) were seen in RA patients but not in RAEB patients. It remains unclear whether lymphocytes in MDS patients were involved in the abnormal (MDS) clones, but our results regarding the increments of CD8+ + CD11a+ and NK cells in RA patients suggest that the mechanism of immune surveillance against the abnormal MDS clones was activated in these RA patients, but not in RAEB patients. Further investigation is required to clarify the functions of these immunoregulatory lymphocytes in MDS patients.     

CD25+ cell depletion hastens the onset of severe disease in collagen‐induced arthritis

Objective

CD4+,CD25+ T regulatory cells may offer opportunities to intervene in the course of autoimmune disease. We wished to evaluate their potential for influencing systemic and chronic joint inflammation by investigating their involvement in collagen‐induced arthritis (CIA).

Methods

We depleted DBA/1 mice of CD25+ regulatory cells by injection of a depleting monoclonal antibody specific for CD25 14 days before a single immunization with type II collagen (CII) in Freund's complete adjuvant. CD4+,CD25+ T cells were adoptively transferred to some groups of mice during immunization. Mice were then scored for signs of arthritis, and blood was taken periodically to measure the amounts of CII‐specific antibodies. Splenocytes of treated mice were examined in vitro to determine the effects of depletion on proliferation to CII and control antigens.

Results

CD25+ cell–depleted DBA/1 mice had significantly more severe disease than control mice following collagen immunization. The magnified severity was also accompanied by higher antibody titers against collagen, and in vitro tests showed increased proliferation of collagen‐specific T cells. Adoptively transferring CD4+,CD25+ T cells into depleted mice was shown to reverse the heightened severity. Control mice, which were depleted and immunized with the neoantigen keyhole limpet hemocyanin (KLH), had neither an increased antibody response toward KLH nor an augmented proliferative response, indicating that CD25+ cell depletion preferentially affects immunity against self antigen.

Conclusion

These results establish a link between CD4+,CD25+ regulatory cells and CIA and provide a rationale for investigating CD4+,CD25+ T regulatory cells in the treatment and prevention of arthritis.