老年原发免疫性血小板减少症患者治疗前后T淋巴细胞亚群的临床研究

目的 检测老年原发免疫性血小板减少症(ITP)患者治疗前后T淋巴细胞亚群的动态变化, 探讨其在ITP发生发展中的作用。方法 采用流式细胞术测定ITP患者治疗前后及正常对照组外周血T淋巴细胞亚群的水平。结果 ITP患者治疗前后T淋巴细胞绝对值、CD3⁺、CD4⁺、CD8⁺、CD4⁺CD25⁺T淋巴细胞比例及CD4⁺/CD8⁺比值分别为(0.83±0.16)vs(1.74±0.36)、(71.71±1.07)% vs(72.69±1.35)%、(41.78±0.71)% vs(42.46±1.20)%、(29.67±0.97)% vs(28.56±1.75)%、(8.76±0.56)% vs(9.39±1.26)%、(1.42±0.07)vs(1.49±0.13), CD8⁺T淋巴细胞比例治疗后显著降低, 其余均显著升高, 差异有统计学意义(P<0.05)。结论 T淋巴细胞亚群的异常改变, 破坏自身免疫, 与病情相关, 可指导临床治疗, 并作为评估预后的参考指标。     

Analysis of the roles of dietary protein and calcium in fluoride‐induced changes in T‐lymphocyte subsets in rat

The roles of dietary protein (Pr) and calcium (Ca) levels on the changes in T‐lymphocyte subsets induced by excessive fluoride (F) intake were assessed using rats that were malnourished for 120 days as a model. The CD⁴⁺ and CD⁸⁺ T‐lymphocytes in the spleen tissue were determined by flow cytometry and immunofluorescence assay. The percentages of CD³⁺, CD⁴⁺, and CD⁸⁺ T‐lymphocytes were reduced in the spleen of rats exposed to excessive F, and malnutrition aggravated these changes in the T‐lymphocytes. In addition, the mRNA expression levels of IL‐1β, IL‐2, IL‐6, TNF‐α, and IFN‐γ in the spleen were downregulated significantly. We also reported herein the increased apoptosis ratio following caspase‐9 and caspase‐3 upregulation in the spleen of rats exposed to excessive amount of F. Light and transmisison electron microscopy revealed the irregularly arranged lymphocytes, few lymph nodules and the apoptotic characteristic of lymphocytes, which are caused by the increased expression of caspase. In addition, Pr and Ca supplementation reversed the morphologic and T‐lymphocytic changes in spleen under malnutrition. Taken together, our results revealed an endogenous caspase‐mediated mechanism of regulating the apoptosis of the T‐lymphocyte subsets, as well as the immune‐related cytokine secretion, which reduces the immune function in F‐induced rats. © 2016 Wiley Periodicals, Inc. Environ Toxicol 32: 1587–1595, 2017.     

Modulating effects of nonselective and selective phosphodiesterase inhibitors on lymphocyte subsets and humoral immune response in mice

Phosphodiesterase (PDE) inhibitors can regulate the activity of immune cells by increasing intracellular levels of cyclic nucleotides. The aim of this study was to determine the effects of milrinone, a selective PDE3 inhibitor, sildenafil, a selective PDE5 inhibitor, and aminophylline, a nonselective PDE inhibitor, on lymphocyte subsets and humoral immune response in mice when administered in vivo. Aminophylline (20 mg/kg, i.m.), milrinone (1 mg/kg, i.m.) or sildenafil (1 mg/kg, p.o.) were administered to mice either once or five times at 24 h intervals. Some mice were immunized with a sheep red blood cell (SRBC) suspension administered i.p. either 2 h after the single dose or 2 h after the second of the five doses. In non-immunized mice treated five times with PDE inhibitors, the subsets of T lymphocytes in the thymus and T and B lymphocytes in the spleen and mesenteric lymph nodes were determined 12, 24 or 72 h after the last dose. The humoral immune response was determined on days 4, 7 and 14 after SRBC injection in SRBC-immunized mice treated with PDE inhibitors. A modulating effect of the drugs on lymphocyte subpopulations was observed. The greatest impact was observed in splenocyte subpopulations, and resulted in decreased percentages of B cells (CD19(+)) and increased percentages of T cells (CD3(+), CD4(+), CD8(+)). No effect or slight influence of the drugs on anti-SRBC hemagglutinins was observed, but the number of plaque-forming splenocytes was increased. The drugs under investigation did not show a significant immunosuppressive effect.     

Effects of lipopolysaccharide on T lymphocyte cell subsets and cytokine secretion in mesenteric lymph nodes of mice: Histological and molecular study

Lipololysaccharides (LPS) can disrupt the gut barrier. How dose LPS affects the immune performance of mesenteric lymph nodes? The results showed the hematological parameters significantly changed after LPS treatment. The length of intestinal villus was shortened and the depth of crypts was deepened, especially on the ileum. After LPS treatment 6 h, 12 h, the number of CD3⁺ T cells and CD4/CD8 in the mesenteric lymph nodes of ileum were reduced significantly; the levels of IFN-γ, TNF-ɑ and IL-2 were significantly decreased, and the levels of IL-6 and IL-10 were significantly increased in the ileum. The content of sIgA in the ileum was significantly decreased after LPS treatment 3 h, 6 h and was increased after LPS treatment 12 h. LPS through mesenteric lymph nodes, which induces the immune function reduced and the ileum injured obviously after treatment 6 h. Furthermore, the performance of intestinal immune performance was the lowest after LPS treatment 6 h.     

早期潜伏梅毒病人外周血T淋巴细胞亚群、自然杀伤细胞的变化及意义

目的 探讨早期潜伏梅毒病人外周血T淋巴细胞亚群、自然杀伤细胞（NK）的变化及意义。方法 选取2015年3月至2021年3月河北北方学院附属第一医院诊治的早期潜伏梅毒病人135例为观察组,另选取同期健康体检者103例为对照组,均采用流式细胞仪检测外周血T淋巴细胞亚群（CD3+、CD4+、CD8+、CD4+/CD8+）、NK细胞水平并进行对比。另根据快速血浆反应素环状卡片试验（RPR）初始滴度将观察组病人分为RPR初始滴度≤1∶8组与>1∶8组,对比RPR初始滴度≤1∶8组与>1∶8组外周血T淋巴细胞亚群、NK细胞检测结果,并采用Spearman秩相关性分析法分析外周血T淋巴细胞亚群、NK细胞与RPR初始滴度的相关性。结果 观察组外周血CD3+、CD8+绝对计数及所占比例[（67.79±10.76）%比（64.85±12.95）%、（29.51±5.92）%比（28.09±5.61）%]与对照组比较均差异无统计学意义（P>0.05）,外周血CD4+绝对计数及所占比例[（37.36±7.26）%比（29.73±5.84）%]、CD4+/CD8+所占比例（1.27±0.25比...     

慢性乙型肝炎患者拉米夫定治疗前后SIL-2R、TNF和T淋巴细胞亚群的变化

目的　研究拉米夫定对慢性乙型肝炎患者SIL 2R(可溶性白细胞介素 2受体 )、TNF(肿瘤坏死因子 )和T淋巴细胞亚群的影响。方法　分别用双抗体夹心酶联免疫法测血清SIL 2R、放射免疫法测血清TNF和单克隆抗体免疫技术测定T淋巴细胞亚群 ,比较拉米夫定治疗 35例慢性乙型肝炎患者治疗前后HBVDNA阴转率、ALT复常率、SIL 2R、TNF和T淋巴细胞亚群的变化。结果　治疗后HBVDNA阴转率为 86 .9% ,ALT复常率为 79.8% ,显著高于治疗前 (P <0 .0 1)。SIL 2R、TNF水平显著降低 (P <0 .0 5 )。T淋巴细胞亚群CD 4 、CD 4 /CD 8水平显著高于治疗前 (P <0 .0 1) ,CD 8显著低于治疗前 (P <0 .0 5 )。HBVDNA阴转者T细胞免疫功能优于未转者。结论　拉米夫定可改善慢性乙型肝炎的细胞因子水平和T细胞免疫功能 ,T细胞免疫功能紊乱较轻的患者 ,拉米夫定治疗的效果较好 ,SIL 2R、TNF及CD 4 /CD 8水平可以作为疗效判断的指标。这些指标在一定程度上反映了机体免疫功能状态和肝脏受损的程度 ,具有较高的临床价值     

Phenethyl isothiocyanate promotes immune responses in normal BALB/c mice,inhibits murine leukemia WEHI‐3 cells,and stimulates immunomodulations in vivo

Enhanced cruciferous vegetable consumption is associated with the reduction of cancer incidence as shown in epidemiological studies. Phenethyl isothiocyanate (PEITC), one of the important compounds in cruciferous vegetables, has been shown to induce apoptosis in many types of human cancer cell lines, but there is no available information addressing the effects on normal and leukemia mice in vivo. The purpose of this study is to focus on the in vivo effects of PEITC on immune responses of normal and WEHI‐3 leukemia BALB/c mice in vivo. Influences of PEITC on BALB/c mice after intraperitoneal (i.p.) injection with WEHI‐3 cells and normal mice were investigated. In normal BALB/c mice, PEITC did not affect the body weight when compared to the olive oil treated animals. Moreover, PEITC promoted phagocytosis by macrophages from peripheral blood mononuclear cells (PBMC) and peritoneal cavity, increased the levels of CD11b and Mac‐3, decreased the level of CD19 and promoted natural killer (NK) cell cytotoxic activity, but it did not alter the level of CD3. Also, PEITC enhanced T cell proliferation after concanavalin A (Con A) stimulation. Otherwise, PEITC increased the body weight, but decreased the weight of liver and spleen as compared to the olive oil‐treated WEHI‐3 leukemia mice. PEITC also increased the level of CD19, decreased the levels of CD3 and Mac‐3 rather than influence in the level of CD11b, suggesting that the differentiation of the precursor of macrophages and T cells was inhibited, but the differentiation of the precursor of B cells was promoted in leukemia mice. Furthermore, PEITC enhanced phagocytosis by monocytes and macrophages from PBMC and peritoneal cavity, and also promoted the NK cell cytotoxic activity in comparison with the group of leukemia mice. Based on these observations, the biological properties of PEITC can promote immune responses in normal and WEHI‐3 leukemia mice in vivo. © 2011 Wiley Periodicals, Inc. Environ Toxicol, 2013.     

Safrole suppresses murine myelomonocytic leukemia WEHI‐3 cells in vivo,and stimulates macrophage phagocytosis and natural killer cell cytotoxicity in leukemic mice

Many anticancer drugs are obtained from phytochemicals and natural products. However, some phytochemicals have mutagenic effects. Safrole, a component of Piper betle inflorescence, has been reported to be a carcinogen. We have previously reported that safrole induced apoptosis in human oral cancer cells in vitro and inhibited the human oral tumor xenograft growth in vivo. Until now, there is no information addressing if safrole promotes immune responses in vivo. To evaluate whether safrole modulated immune function, BALB/c mice were intraperitoneally injected with murine myelomonocytic WEHI‐3 leukemia cells to establish leukemia and then were treated with or without safrole at 4 and 16 mg/kg. Animals were sacrificed after 2 weeks post‐treatment with safrole for examining the immune cell populations, phagocytosis of macrophages and the natural killer (NK) cells' cytotoxicity. Results indicated that safrole increased the body weight, and decreased the weights of spleen and liver in leukemic mice. Furthermore, safrole promoted the activities of macrophages phagocytosis and NK cells' cytotoxicity in leukemic mice when compared with untreated leukemic mice. After determining the cell marker population, we found that safrole promoted the levels of CD3 (T cells), CD19 (B cells) and Mac‐3 (macrophages), but it did not affect CD11b (monocytes) in leukemic mice. In conclusion, safrole altered the immune modulation and inhibited the leukemia WEHI‐3 cells in vivo. © 2011 Wiley Periodicals, Inc. Environ Toxicol 28:601–608, 2013.     

4‐fluorotranylcypromine,a novel monoamine oxidase inhibitor: Neurochemical effects in rat brain after short‐ and long‐term administration

A series of acute and chronic experiments was conducted on 4‐fluorotranylcypromine (FTCP), an analog of the monoamine oxidase (MAO)‐inhibiting antidepressant tranylcypromine (TCP) in which the 4 position of the phenyl ring is protected from metabolism. These studies demonstrated that FTCP is a more potent inhibitor of MAO ex vivo than is the parent drug. Despite its similarity in structure to p‐chloroamphetamine, FTCP does not cause a depletion of brain levels of 5‐hydroxytryptamine (5‐HT) after chronic administration; in fact, it increases brain 5‐HT to levels similar to those produced by TCP. After chronic administration, FTCP produces a downregulation of β‐adrenergic and tryptamine receptors, in common with TCP and several other antidepressants. Pretreatment of rats with iprindole or trifluperazine, drugs known to block cytochrome P450‐mediated hydroxylation reactions and to elevate brain levels of TCP, had no effects on FTCP brain levels, suggesting that metabolic drug–drug interactions may be less of a concern with FTCP than with TCP. In vitro uptake experiments in prisms prepared from hypothalamus or striatum revealed that TCP and FTCP are both potent inhibitors of norepinephrine (NE) uptake; FTCP is a more potent inhibitor of 5‐HT uptake than is TCP. FTCP is a more potent releaser of 5‐HT than is TCP. Drug Dev. Res. 48:61–69, 1999. © 1999 Wiley‐Liss, Inc.     

Increases in adipose and total body weight,but not in lean body mass,associated with subcutaneous administration of Sonic hedgehog‐Ig fusion protein to mice

Sonic hedgehog (Shh) is a member of a family of proteins that are involved in embryonic development. The receptor signaling pathways for Shh persist in adults and stimulation of this pathway has shown therapeutic efficacy in animal models of neurodegeneration/regeneration. This study was conducted to evaluate the safety of repeat dose administration of an IgG fusion protein of Shh (Shh‐Ig) in adult mice. Routine toxicology evaluations were performed. In addition, body composition analysis was conducted by densitometry. Shh‐Ig treatment caused a significant increase in body weight gain relative to controls and a slight increase in liver and spleen weights. The increase in body weight could be largely accounted for by an increase in body fat. The treatment‐related increases in body weight were reversible upon cessation of treatment. Shh‐Ig treatment produced no significant changes in clinical chemistry or hematology. There were no gross or histomorphologic findings in any tissue except for skin and spleen. Microscopic findings in the skin were limited to minimal to slight local epidermal hyperplasia at the sc injection site and increased thickness of the fat layer. In the spleen a slight increase in extramedullary hematopoiesis was seen. This finding is possibly a secondary response following inflammation at the injection site in some animals due to the administration of a foreign protein. This study showed that Shh‐Ig administration was well tolerated. The most significant finding was a reversible increase in body weight. Drug Dev. Res. 57:107–114, 2002. © 2002 Wiley‐Liss, Inc.     

Effects of diallyl trisulfide on induction of apoptotic death in murine leukemia WEHI‐3 cells in vitro and alterations of the immune responses in normal and leukemic mice in vivo

Diallyl trisulfide (DATS), a chemopreventive dietary constituent and extracted from garlic, has been shown to against cultured many types of human cancer cell liens but the fate of apoptosis in murine leukemia cells in vitro and immune responses in leukemic mice remain elusive. Herein, we clarified the actions of DATS on growth inhibition of murine leukemia WEHI‐3 cells in vitro and used WEHI‐3 cells to generate leukemic mice in vivo, following to investigate the effects of DATS in animal model. In in vitro study, DATS induced apoptosis of WEHI‐3 cells through the G0/G1 phase arrest and induction of caspase‐3 activation. In in vivo study DATS decreased the weight of spleen of leukemia mice but did not affect the spleen weight of normal mice. DATS promoted the immune responses such as promotions of the macrophage phagocytosis and NK cell activities in WEHI‐3 leukemic and normal mice. However, DATS only promotes NK cell activities in normal mice. DATS increases the surface markers of CD11b and Mac‐3 in leukemia mice but only promoted CD3 in normal mice. In conclusion, the present study indicates that DATS induces cell death through induction of apoptosis in mice leukemia WHEI‐3 cells. DATS also promotes immune responses in leukemia and normal mice in vivo. © 2014 Wiley Periodicals, Inc. Environ Toxicol 30: 1343–1353, 2015.     

Toxicological evaluation of the topoisomerase inhibitor,etoposide, in the model animal Caenorhabditis elegans and 3T3‐L1 normal murine cells

Etoposide, a topoisomerase II inhibitor, has been widely used as a clinical anticancer drug to treat diverse cancer patients. Since not only rapidly dividing cancer cells but also the cells of normal human tissues and every living organism in environmental ecosystems have topoisomerases, it is crucial to study the toxicity of etoposide in other organisms in addition to cancer cells. In this study, we evaluated the toxicity of etoposide in both a soil nematode, Caenorhabditis elegans, and 3T3‐L1 normal murine cells. Etoposide significantly retarded the growth, egg laying, and hatching in C. elegans. Etoposide also affected the reproductive gonad tissue, decreased the number of germ cells and induced abnormally enlarged nuclei in C. elegans. In addition, etoposide inhibited 3T3‐L1 cell proliferation, with IC₅₀ values of 37.8 ± 7.3 and 9.8 ± 1.8 μM after 24 and 48 hours of treatment, respectively, via the induction of cell cycle arrest at the G2/M phase and apoptotic cell death. Etoposide also induced nuclear enlargement in 3T3‐L1 normal murine cells. The reproductive toxicity and abnormal nuclear morphological changes seemed to correlate with the adverse effects of etoposide. We suggest that these experimental platforms, i.e., the toxicological evaluation of both nematodes and 3T3‐L1 cells, may be useful to study the mechanisms underlying the side effects of chemicals, including topoisomerase inhibitors.     

Melatonin attenuates 60Co γ‐ray‐induced hematopoietic,immunological and gastrointestinal injuries in C57BL/6 male mice

Protection of hematopoietic, immunological, and gastrointestinal injuries from deleterious effects of ionizing radiation is prime rational for developing radioprotector. The objective of this study, therefore, was to evaluate the radioprotective potential of melatonin against damaging effects of radiation‐induced hematopoietic, immunological, and gastrointestinal injuries in mice. C57BL/6 male mice were intraperitoneally administered with melatonin (50–150 mg/kg) 30 min prior to whole‐body radiation exposure of 5 and 7.5 Gy using ⁶⁰Co‐teletherapy unit. Thirty‐day survival against 7.5 Gy was monitored. Melatonin (100 mg/kg) pretreatment showed 100% survival against 7.5 Gy radiation dose. Melatonin pretreatment expanded femoral HPSCs, and inhibited spleenocyte DNA strands breaks and apoptosis in irradiated mice. At this time, it also protected radiation‐induced loss of T cell sub‐populations in spleen. In addition, melatonin pretreatment enhanced crypts regeneration and increased villi number and length in irradiated mice. Translocation of gut bacteria to spleen, liver and kidney were controlled in irradiated mice pretreated with melatonin. Radiation‐induced gastrointestinal DNA strand breaks, lipid peroxidation, and expression of proapoptotic‐p53, Bax, and antiapoptotic‐Bcl‐xL proteins were reversed in melatonin pretreated mice. This increase of Bcl‐xL was associated with the decrease of Bax/Bcl‐xL ratio. ABTS and DPPH radical assays revealed that melatonin treatment alleviated total antioxidant capacity in hematopoietic and gastrointestinal tissues. Present study demonstrated that melatonin pretreatment was able to prevent hematopoietic, immunological, and gastrointestinal radiation‐induced injury, therefore, overcoming lethality in mice. These results suggest potential of melatonin in developing radioprotector for protection of bone marrow, spleen, and gastrointestine in planned radiation exposure scenarios including radiotherapy. © 2016 Wiley Periodicals, Inc. Environ Toxicol 32: 501–518, 2017.     

Influence of cyclophosphamide and its metabolic products on the activity of peritoneal macrophages in mice

2,4,6-Trinitrophenyl (TNP) hapten-labeled peritoneal macrophages (Mf) given intravenously (iv) to recipients are poor inducers of contact sensitivity (CS) reactions unless Mf donors are pretreated with low doses of cyclophosphamide (CY). In vivo CY is converted into active alkylating metabolites, phosphoramide mustard (PM) and acrolein (ACR).Our experiments aimed to test how in vitro treatment of non-immunogenic Mf with different concentrations (10^?5 to 10^?7 M) of CY metabolites will influence their immunogenicity and other biological functions. Instead of chemically unstable PM, we used structurally and functionally similar nitrogen mustard (NM).Our experiments show that treatment of Mf with ACR or NM stimulates the in vitro production of pro-inflammatory IL-6 and IL-12 and down-regulates anti-inflammatory IL-10 and TGF-β cytokines. In vivo non-immunogenic TNP-Mf become capable of inducing CS reactions in two situations: first, after treatment with NM or ACR and second, when cell recipients are received iv before Mf transfer of monoclonal antibodies against IL-10 and/or TGF-β (500 μg per animal). Treatment with NM, but not with ACR, was also an efficient stimulus for production by Mf of significantly increased levels of reactive oxygen intermediates (ROIs).In summary, our experiments show that CY metabolites can significantly increase the specific immune response as well as nonspecific innate reaction (ROIs production) and support the notion that CYand its metabolites can be a promising accessory tool when upregulation of the immune response is desired.