Reconstitución de células natural killer después del trasplante de células progenitoras hematopoyéticas en niños / Natural killer cell reconstitution after hematopoietic stem-cell transplantation in children

Resumen Introducción: Después de un trasplante de células progenitoras hematopoyéticas (TCPH), la reconstitución de las células natural killer (NK) es la principal barrera contra las infecciones virales. Objetivo: Determinar que el conocimiento sobre la cinética de la reconstitución de las células NK posterior al TCPH contribuye a un eficiente monitoreo del trasplante, lo que incrementa la posibilidad de éxito de este. Método: Se incluyeron 21 pacientes sometidos a TCPH, así como un grupo control de individuos clínicamente sanos. En diferentes momentos después del trasplante (intervalo de 21 a 670 días), mediante citometría de flujo se cuantificaron las células NK CD3− CD16+ CD56+ en muestras de sangre periférica. Resultados: La recuperación de las células NK ocurre entre los tres y seis meses y entre los 10 y 12 meses postrasplante; su número fue significativamente menor (en comparación con el grupo control) en el tiempo restante del monitoreo. Conclusiones: El primer periodo de recuperación de las células NK ocurre entre los tres y seis meses posteriores al trasplante. La reconstitución es transitoria y el número de células NK varía en los primeros años.

Abstract Introduction: After hematopoietic stem cell transplantation (HSCT), natural killer (NK) cells reconstitution is the main barrier against viral infections. Objective: To determine that the knowledge on the kinetics of NK cell reconstitution after HSCT contributes to transplant efficient monitoring, which increases the possibility of its success. Method: Twenty-one patients undergoing HSCT were included, as well as a control group of clinically healthy individuals. At different time points after transplantation (range of 21 to 670 days), CD3- CD16+ CD56+ NK cells were quantified by flow cytometry in peripheral blood samples. Results: NK cell recovery occurs at three to six months and 10 to 12 months post-transplantation; their number was significantly lower (in comparison with the control group) in the rest of the monitoring time. Conclusions: The first period of NK cell recovery occurs between three and six months after transplantation. Reconstitution is transient and the number of NK cells varies in the first years.

Subsets of T lymphocytes in the lesional skin of pityriasis rosea

Abstract: Background: Pityriasis rosea is a common papulosquamous disorder. However, its etiology and pathogenesis remain unclear. Objective: We investigate the types of inflammatory cells infiltrating the lesional skin of pityriasis rosea and demonstrate whether T-cell-mediated immunity is involved in the pathogenesis of this condition or not. Methods: The biopsies were taken from the lesional skin of 35 cases of patients diagnosed with pityriasis rosea. The specimens were prepared in paraffin sections, then submitted to routine immunohistochemistry procedures using monoclonal antibodies directed against CD3, CD4, CD8, CD20 and CD45RO and horseradish peroxidase-labeled goat anti-human antibodies. The positive sections were determined by the ratio and staining intensity of positive inflammatory cells. Results: The mean score of positive CD3, CD4, CD8, and CD45RO staining was respectively 3.74±3.88, 5.67±4.40, 2.94±3.42 and 7.68±4.33 in these pityriasis rosea patients (P<0.001). The percentage of positive staining was 54.29% (19/35), 69.7% (23/33), 40% (14/35) and 79.41% (27/34) (P<0.05). However, the staining of CD20 was negative in all samples. The mean score of CD3 staining in patients with time for remission ≤60 days (4.90±4.21) was higher than that in patients with time for remission >60 days (2.00±2.5) (P<0.05), whereas no statistical difference in the mean score of CD4, CD8 and CD45RO staining was observed. study liMitations: The sample size and the selected monoclonal antibody are limited, so the results reflect only part of the cellular immunity in the pathogenesis of pityriasis rosea. Conclusion: Our findings support a predominantly T-cell mediated immunity in the development of pityriasis rosea.

Effects of baicalin on inflammatory reaction, oxidative stress and PKDl and NF-kB protein expressions in rats with severe acute pancreatitis

Abstract Purpose: To investigate the effects of baicalin on inflammatory reaction, oxidative stress and protein kinase D1 (PKD1) and nuclear factor-kappa B (NF-κB) protein expressions in severe acute pancreatitis (SAP) rats. Methods: Sixty rats were divided into sham operation, model, and low-, medium- and high-dose baicalin group. SAP model was established in later 4 groups. The later 3 groups were injected with 0.1, 0.2 and 0.4 ml/100 g 5% baicalin injection, respectively. At 12 h, the serum SAP related indexes and inflammatory factors, peripheral blood CD3 and γδT cell percentages, wet/dry ratio and pancreas ascites volume, oxidative stress indexes and PKD1 and NF-κB protein expressions in pancreatic tissue were determined. Results: Compared with model group, in high-dose baicalin group the wet/dry ratio and ascites volume, serum amylase level, phospholipase A2 activity, TNF-α, IL-1 and IL-6 levels, and pancreatic malondialdehyde level and PKD1 and NF-κB protein expression were significantly decreased (P < 0.05), and peripheral blood CD3 and γδT cell percentages and pancreatic superoxide dismutase and glutathione peroxidase levels were significantly increased (P < 0.05). Conclusion: Baicalin can resist the inflammatory reaction and oxidative stress, and down-regulate protein kinase D1 and nuclear factor-kappa B protein expressions, thus exerting the protective effects on severe acute pancreatitis in rats.

Potent and conditional redirected T cell killing of tumor cells using Half DVD-Ig

Novel biologics that redirect cytotoxic T lymphocytes (CTLs) to kill tumor cells bearing a tumor associated antigen hold great promise in the clinic. However, the ability to safely and potently target CD3 on CTL toward tumor associated antigens (TAA) expressed on tumor cells remains a challenge of both technology and biology. Herein we describe the use of a Half DVD-Ig format that can redirect CTL to kill tumor cells. Notably, Half DVD-Ig molecules that are monovalent for each specificity demonstrated reduced non-specific CTL activation and conditional CTL activation upon binding to TAA compared to intact tetravalent DVD-Ig molecules that are bivalent for each specificity, while maintaining good drug like properties and appropriate PK properties.

System Amplifying NK Cells Using Mononuclear Cell Culture / 中国实验血液学杂志

<p><b>OBJECTIVE</b>To develop an easy method to amplify natural killer (NK) cells by using mononuclear cells in vitro, so as to lay the basis for NK cell therapy.</p><p><b>METHODS</b>Umbilical cord blood from 3 healthy full-term pregnant women was collected, and the peripheral blood mononuclear cells (PBMNC) were harvested by density gradient centrifugation. Each sample of PBMNC was divided into 3 groups: CD16mAb, CD3 mAb and CD16mAb+CD3mAb- groups. The culture flasks were pre-coated with CD16, CD3 or CD3 plus CD16 mAb. The PBMNCs were cultured in serum-free media containing autologous plasma, recombinant human IL-2, IL-15 and IL-21 for 14 days under the same conditions. The total viable cell count was calculated. Flow cytometry was used to determine the ratio of CD56CD3 cells, MTT assay was used to measure the killing rate of NK cells under different effector/target ratio, by using the K562 cells as the target cells.</p><p><b>RESULTS</b>After 14 days of culture, the total cell numbers of CD16mAb, CD3mAb and CD16mAb +CD3mAb groups increased by 45.71±5.54, 87.41±19.77 and 4.88±51.84 times, respectively, and those of CD3mAb group were significantly higher than the other 2 groups (P<0.05). The ratio of CD56CD3 cells before culture was 0.1663±0.0201, which was 0.8167±0.0500, 0.8077±0.0589 and 0.8077±0.0273 after incubation with CD16mAb, CD3mAb and CD16mAb +CD3mAb for 14 days, respectively (P>0.05). MTT test showed that the killing efficiencies were not significantly different among the 3 groups when the effector/target ratios were 1:1, 5:1 and 10:1 (P>0.05).</p><p><b>CONCLUSION</b>By incubation with anti-CD3 monoclonal antibody, IL-2, IL-15 and IL-21, the highly purified NK cells can be obtained from mononucleated cells, thus providing a simple method for NK cell therapy.</p>

Polyclonal anti T-lymphocyte antibody therapy monitoring in kidney transplant recipients: comparison of CD3+ T cell and total lymphocyte counts / Monitoramento da terapia com anticorpo policlonal antilinfócitos T em transplantados renais: comparação da contagem de células T CD3+ e de linfócitos totais

ABSTRACT Objective: To investigate the correlation between total lymphocyte and CD3+ T cell counts in peripheral blood in renal transplant patients treated with anti-thymocyte globulin, and discuss related outcomes. Methods: A single-center, retrospective study involving 226 patients submitted to kidney transplant between 2008 and 2013, and treated with anti-thymocyte globulin for induction or treatment of cellular rejection. Doses were adjusted according to CD3+ T cell or total lymphocyte counts in peripheral blood. Results: A total of 664 paired samples were analyzed. The Spearman's correlation coefficient was 0.416 (p<0.001) for all samples combined; the overall Kappa coefficient was 0.267 (p<0.001). Diagnostic parameters estimated based on total lymphocyte counts were also calculated using the number of CD3+ T cells (gold standard), with a cut off of >20 cells/mm3. Conclusion: Total lymphocyte and CD3+ T cell counts in peripheral blood are not equivalent monitoring strategies in anti-thymocyte globulin therapy.

RESUMO Objetivo: Investigar a correlação entre a contagem de linfócitos totais e células T CD3+ no sangue periférico em receptores de transplante renal submetidos a tratamento com globulina antitimocitária, e discutir resultados relacionados. Métodos: Estudo retrospectivo de centro único envolvendo 226 pacientes submetidos a transplante renal entre 2008 e 2013 e tratados com globulina antitimocitária, para fins de indução ou tratamento de rejeição celular. As doses foram ajustadas de acordo com a contagem de células T CD3+ ou linfócitos totais no sangue periférico. Resultados: No total, 664 amostras pareadas foram analisadas. O coeficiente de correlação de Spearman para as amostras em geral foi de 0,416 (p<0,001) e o coeficiente Kappa, de 0,267 (p<0,001). Os parâmetros diagnósticos estimados com base na contagem de linfócitos totais foram recalculados, empregando-se o número de células T CD3+ (padrão-ouro) e adotando-se o ponto de corte >20 células/mm3. Conclusão: A contagem de linfócitos totais no sangue periférico não substitui a contagem de células T CD3+ enquanto estratégia de monitorização da terapia à base de globulina antitimocitária.

Cut-off value for absolute lymphocytes as an alternative for the immunophenotypic analysis of CD3+ T cells in the monitoring of immunosuppressive therapy with thymoglobulin / Valor de corte para linfócitos absolutos como alternativa para análise imunofenotípica de células T CD3+ no monitoramento de terapia imunossupressora com timoglobulina

Abstract Introduction: Immunosuppression of T lymphocytes is required for preventing acute rejection after transplantation and for the treatment of chronic autoimmune and inflammatory diseases. The laboratory monitoring for this therapy is the measurement of T cells by immunophenotyping, aiming the target value of less than 20 cells per µL. Objective: To establish a cut-off point for the total number of lymphocytes in the automated blood cell count that reflects less than twenty T cells µL by immunophenotyping. Methods: We studied and evaluated 242 kidney transplant patients that had results of automated blood cell count and quantification of T cells by immunophenotyping technique. The patients were divided into two groups, depending on the T lymphocyte immunophenotyping rates established by lower and higher than 20 cells per µL. After, we evaluated the cut-off point for lymphocytes in the blood cell count with a specificity of 100% to exclude patients with high levels of T lymphocytes. Results: We found that the cut-off point of 70 lymphocytes per µL obtained by automated blood cell count showed 100% of specificity to exclude patients with T-cell counts higher than 20 cells per µL by immunophenotyping. Conclusion: The results found in this study may be helpful to monitor the immunosuppressive therapy in kidney transplant patients in places where a flow cytometer is not available, or when this equipment is not present in the full routine.

Resumo Introdução: A imunossupressão de linfócitos T é necessária para a prevenção da rejeição aguda em transplantes e no tratamento de doenças autoimunes e inflamatórias crônicas. O seu monitoramento laboratorial consiste na quantificação dos linfócitos T realizada pela técnica de imunofenotipagem, na qual o valor preconizado é manter inferior a 20 células/µL. Objetivo: Estabelecer um ponto de corte para o número de linfócitos totais no hemograma automatizado que reflita uma contagem de linfócitos T inferior a 20 células/µL por imunofenotipagem. Métodos: Foram avaliados 242 pacientes transplantados renais que continham resultados do hemograma automatizado e quantificação de linfócitos T por imunofenotipagem. Os pacientes foram divididos em dois grupos, conforme os valores de linfócitos T estabelecidos pela imunofenotipagem: inferiores e superiores a 20 células/µL. A partir disto, foi avaliado o ponto de corte de linfócitos no hemograma com especificidade de 100% para excluir os pacientes com valores elevados de linfócitos T. Resultados: Este estudo evidenciou que o ponto de corte de 70 linfócitos/µL obtidos pelo hemograma automatizado apresentou especificidade de 100% para excluir os pacientes com contagens de linfócitos T superiores a 20 células/µL na imunofenotipagem. Conclusão: Esta pesquisa poderá auxiliar no monitoramento da terapia imunossupressora em pacientes transplantados renais em locais que não possuem um citômetro de fluxo disponível, ou ainda quando este equipamento não se faz presente na rotina integral.

IL32 expression in peripheral blood CD3+ cells from myelodysplastic syndromes patients

Background: Myelodysplastic syndromes (MDS) are a heterogeneous group of disorders characterized by ineffective hematopoiesis and risk of leukemia transformation. There is evidence to suggest the participation of immune system deregulation in MDS pathogenesis. Interleukin-32 (IL-32) is a newly described multifunctional cytokine reported as an important mediator in autoimmune and inflammatory disorders. In the present study, we reported the expression of IL32 and IL32 transcript variants (α, ß, γ and δ) in peripheral blood CD3+ cells from healthy controls and MDS patients. Methods: CD3+ cells were isolated by immunomagnetic cell sorting from thirty-nine untreated MDS patients and twenty-nine healthy donors. Gene expression was evaluated by quantitative PCR. For statistical analysis, Mann­Whitney test, Kruskal-Wallis test with Dunns post test and Log-rank (Mantel-Cox) were used, as appropriate. A p value <0.05 was considered statistically significant. Results: IL32 expression and IL32 transcript variants IL32α, IL32ß, IL32γ, and IL32δ, were similar in peripheral blood CD3+ cells from healthy donors and MDS patients. Increased IL-32α expression was an independent predictor for MDS disease progression by univariate and multivariate analysis. Conclusions: We observed that IL32 expression is not differently expressed in CD3+ cells from MDS patients; nevertheless IL32α has a potential role in disease progression (AU)

Effects of extracellular heat shock protein 70 on intestinal immune function of rats with severe scald injury / 中华烧伤杂志

<p><b>OBJECTIVE</b>To explore the change in the expression of extracellular heat shock protein 70 (eHSP70) and interleukin 2 (IL-2) and their correlation in intestine of rats with severe scald injury, and to observe the effects of eHSP70 on CD3(+) T lymphocytes in Peyer's patch of intestine in rats with severe scald injury in vitro.</p><p><b>METHODS</b>(1) Sixty male SD rats were divided into normal control group (NC, n=10, only anesthetized) and scald group (S, n=50) according to the random number table. Rats in scald group were inflicted with 30% total body surface area full-thickness scald on the back. Ten rats from group NC immediately after anesthetization and 10 rats from group S at post injury hour (PIH) 3, 6, 12, 24, 48 were sacrificed to harvest their small intestines. The expressions of eHSP70 and IL-2 were determined with enzyme-linked immunosorbent assay (ELISA), and their correlation was analyzed. (2) Another 2 male SD rats were inflicted with the same injury as above. At PIH 12, CD3(+) T lymphocytes in Peyer's patch of small intestine were isolated and cultured with RPMI 1640 nutrient solution containing 10% fetal bovine serum. Cells were divided into blank control group (BC) and 5, 10, 20 μg/mL eHSP70 groups according to the random number table, with 6 wells in each group. Cells in group BC didn't receive any other treatment, while cells in the latter three groups were treated with corresponding mass concentration of recombinant rat eHSP70. After being cultured for 48 hours, the proportions of Th1 and Th2 in CD3(+) T lymphocytes, and the apoptosis rate of CD3(+) T lymphocytes were detected with flow cytometer, while the expressions of IL-2 and IL-10 in culture supernatant of cells were determined with ELISA. The cell experiments were repeated for 10 times. Data were processed with one-way analysis of variance, Kruskal-Wallis rank sum test, SNK-q test, and Pearson correlation analysis.</p><p><b>RESULTS</b>(1) Compared with those in group NC [(1 278±135) and (48.6±4.9) ng/mg], the levels of eHSP70 [(728±93), (412±31), (314±21), (528±40), (1 028±97) ng/mg] and IL-2 [(38.6±2.3), (32.3±1.0), (25.3±3.6), (33.9±4.1), (44.3±2.6) ng/mg] in intestine of rats in group S obviously decreased at PIH 3, 6, 12, 24, 48 (with q values from 3.48 to 5.32, P values below 0.05), reaching the nadir both at PIH 12, with a significantly positive correlation between the level of IL-2 and the level of eHSP70 (r=0.920, P<0.01). (2) Compared with those in group BC [(8.6±1.1)% and (3.75±0.45)%], the proportion of Th1 obviously increased [(11.3±2.1)%, (15.7±1.8)%, (10.8±1.5)%, with q values from 2.97 to 4.57, P values below 0.05], while the proportion of Th2 obviously decreased [(2.39±0.38)%, (1.05±0.23)%, (2.67±0.26)%, with q values from 2.48 to 4.32, P values below 0.05] in CD3(+) T lymphocytes of rats in 5, 10, 20 μg/mL eHSP70 groups. Compared with those in group BC [(34.3±2.2)% and (254±16) pg/mL], the apoptosis rate of CD3(+) T lymphocytes obviously decreased [(26.1±2.6)%, (20.7±1.5)%, (31.5±2.4)%, with q values from 3.47 to 4.95, P values below 0.05], while the level of IL-2 obviously increased [(417±22), (587±19), (307±27) pg/mL, with q values from 3.02 to 4.98, P values below 0.05] in culture supernatant of CD3(+) T lymphocytes of rats in 5, 10, 20 μg/mL eHSP70 groups. There was no significant difference in the level of IL-10 in culture supernatant of CD3(+) T lymphocytes of rats among the four groups (F=2.12, P>0.05).</p><p><b>CONCLUSIONS</b>The expressions of eHSP70 and IL-2 in intestine of rats are decreased after severe scald, with a obviously positive correlation between them. eHSP70 can promote the differentiation of CD3(+) T lymphocytes in Th1 orientation, decrease the apoptosis rate of the cells, and promote the release of IL-2 of cells in Peyer's patch of intestine in rats with severe scald injury in vitro.</p>

Qingre quyu granule stabilizes plaques through inhibiting the expression of tenascin-C in patients with severe carotid stenosis / 中国结合医学杂志

<p><b>OBJECTIVE</b>To investigate the therapeutic effects of Qingre Quyu Granule (QQG) on the patients with severe carotid stenosis, and to explore the mechanism of it.</p><p><b>METHODS</b>Ninety-six patients with severe carotid stenosis were enrolled in the study and were classified into a QQG group (n=48) and a control group (n=48) randomly using consecutively numbered envelopes. The patients in the QQG group were given QQG and Western medicine, those in the control group were given Western medicine merely, the course of treatment was 16 weeks. All patients went through endarterectomy after treatment. Plaques were subjected to the analysis of CD3, CD68, soluble intercellular adhesion molecule 1 (ICAM-1), matrix metalloprotease-9 (MMP-9), CD40L, tenascin-C, and collagen content lipid content by immunohistochemistry or polarized light analysis.</p><p><b>RESULTS</b>By the end of experiment, the expressions of CD3, CD68, ICAM-1, MMP9, CD40L and tenascin-C on the plaques were statistically significant lower in the QQG group compared with the control group(P<0.01). The lipid content of the plaque was also significantly lower in the QQG group compared with the control group (P<0.01). The interstitial collagen in the tissue sections of the plaques was also significantly higher in the QQG group in comparison with the control group (P<0.01).</p><p><b>CONCLUSION</b>QQG could stabilize carotid artery plaques through inhibiting pro-inflammation factors and restraining the tenascin-C and MMP9 pathway.</p>

Prognostic significance of tumor-infiltrating CD8⁺ or CD3⁺ T lymphocytes and interleukin-2 expression in radically resected non-small cell lung cancer / 中华医学杂志(英文版)

<p><b>BACKGROUND</b>Altered immunoresponse is associated with tumorigenesis and cancer progression. This study assessed the levels of tumor-infiltrating CD3 + or CD8 + T lymphocytes and interleukin-2 (IL-2) protein in radically resected non-small cell lung cancer (NSCLC) tissues to predict overall survival (OS) of the patients.</p><p><b>METHODS</b>Paraffin-embedded tissue specimens from 129 NSCLC patients were retrospectively collected for immunostaining of CD8 + , CD3 + , and IL-2 expression. Clinicopathological and survival data were collected and analyzed using the Chi-squared test, Kaplan-Meier curves, and the log-rank test or the Cox regression model.</p><p><b>RESULTS</b>The data showed a significant inverse association between CD8 + T lymphocyte levels and IL-2 expression (r = -0.927; P = 0.000) and between the levels of CD8 + and CD3 + T lymphocytes (r = -0.722; P = 0.000), but a positive association between CD3 + T lymphocyte levels and IL-2 expression (r = 0.781; P = 0.000) in NSCLC tissues. Furthermore, the levels of CD3 + and CD8 + T lymphocytes and IL-2 expression were associated with tumor stage (P = 0.023, 0.006, and 0.031, respectively) and the level of CD8 + T lymphocytes was associated with the patient gender (P = 0.024). In addition, the levels of CD8 + T lymphocytes were associated with an unfavorable 5-year OS, whereas patients with high levels of CD3 + T lymphocytes in tumor lesions and IL-2-expressing tumors had significantly better 5-year OS rates than patients with low levels.</p><p><b>CONCLUSIONS</b>The levels of CD8 + T cells in tumor lesions and IL-2 expression were both independent predictors of OS for these NSCLC patients. Thus, the detection of tumor-infiltrating CD3 + or CD8 + T lymphocytes and IL-2 expression could be useful to predict the prognosis of radically resected NSCLC patients.</p>

Peptidic HIV-1 fusion inhibitor VIR576 as a potential dual- functional microbicide inhibits antigen-specific CD4(+) T-cell activation / 南方医科大学学报

<p><b>OBJECTIVE</b>To observe if VIR576, an 20-mer peptide derived from the C-proximal subfragment of a1-antitrypsin (a1-AT) which inhibits human immunodeficiency virus type 1 (HIV-1) entry into the target cells by interacting with fusion peptide (FP), can also directly inhibit CD4(+) T cell activation in vitro.</p><p><b>METHODS</b>Splenocytes isolated from DO11.10 OVA Tg mice were stimulated with ovalbumin or concanavalin A to test the effects of VIR576 on antigen-specific or non-antigen-specific T cell activation. Both primary CD4(+)CD25(-) T cells from DO11.10 mice and CD4(+) T cell line A2b were activated with specific antigens to evaluate the effects of VIR576.</p><p><b>RESULTS</b>VIR576 inhibited antigen-specific splenocyte activation but had no significant effect on non-antigen-specific T-cell activation, which bypassed the crosstalk between the CD3-signaling complex and TCR. We furthermore observed that VIR576 could also down-regulate antigen-specific CD4(+) T-cell activation.</p><p><b>CONCLUSIONS</b>Given the high susceptibility of activated CD4(+) T cells in the mucosa to HIV-1 infection, the inhibitory effects of VIR576 on both HIV entry into the target cells and CD4(+) T-cell activation suggest the potential of VIR576 as a microbicide for prevention of sexual transmission of HIV.</p>

Immunophenotyping of bone marrow biopsies in 26 patients with non-hodgkin's lymphoma, using anti-CD3, CD8, CD19 and CD20 monoclonal antibodies

Immunophenotyping with monoclonal antibodies [MoAbs] directed against lymphoid-associated antigens, immunohistochemical staining on paraffin-embedded BM biopsy material, and molecular studies of Ig genes/T-cell receptor genes or lymphoma-associated gene translocations should be used in the global approach to the patient with malignant lymphoma. 1. To determine the subtypes of non-Hodgkin lymphomas [B- or T-cell] in the bone marrow using anti-CD3, CD8 monoclonal antibodies for T-cell and anti-CD19 and CD20 for B-cells. 2. Correlation of the subtypes of NHL [B- or T-cell] with the morphology and pattern of bone marrow infiltration. A retrospective study, done in Al-Kadhymia teaching hospital during the period from 1/10/2010 to1/2/2011.The study consisted 26 adult patients, who were diagnosed as Non-Hodgkin lymphomas by undergoing a BM biopsy. Immunohistochemical staining of the paraffin-embedded sections of BM trephine biopsies was performed in all cases and used standard techniques with monoclonal anti-CD8, CD20 and dual immunofluorscence-labelled CD3, and CD19 antibodies and also all stained with Hematoxylin and Eosin [H and E] for morphologic assessment. The 26 cases of NHL comprised of 14 male [54%] and 12 female patients [46%]. The median age was [57.32] year ranged from 27-85 years. There were 23 cases of B-cell cases [88.5%] and 3 cases of T-cell lineage [11.5%] of all the cases. Among all the B-cell lymphomas, 15 cases showed interstitial infiltration in the bone marrow, while among the T-cell lymphoma two cases showed diffuse infiltration. 1. In these 26 cases NHL patients with marrow involvement, B cell phenotype comprised 88% of cases. 2. B-cell NHLs had predominance of interstitial infiltration in bone marrow biopsies in comparison with the T-cell lymphoma, in which diffuse infiltration was predominant

Presence of CD3+ tumor infiltrating lymphocytes is significantly associated with good prognosis in infiltrating ductal carcinoma of breast.

BACKGROUND: Aim of this study was to investigate the prognostic significance of CD3+ TILs in infiltrating ductal carcinoma (IDC) of the breast. MATERIALS AND METHODS: Immuno-histochemistry was done with CD3 antibodies in tissue sections of 127 breast cancer patients, and CD3+ intra-tumoral and stromal TILs were counted in relation to clinico-pathological variables. RESULTS: Intra-tumoral and stromal CD3+ TILs were significantly associated with positive lymph node status (P = 0.006, P = 0.043, respectively) without significant association with age, menopausal status, family history, and hormonal status. The higher CD3 intra-tumoral and stromal counts both showed significant association with good prognosis (P = 0.039, P = 0.044, respectively). The intra-tumoral count was higher than stromal count and was independently associated with disease-free survival in stage I and II cancer (P = 0.021). CONCLUSIONS: CD3+ TILs may serve as independent marker of good prognosis in IDC breast. The findings of this study need further validation on a larger sample size.

Reactive lymphoid hyperplasia of the liver

No abstract available.

Antiviral effect of dietary germanium biotite supplementation in pigs experimentally infected with porcine reproductive and respiratory syndrome virus

Germanium biotite (GB) is an aluminosilicate mineral containing 36 ppm germanium. The present study was conducted to better understand the effects of GB on immune responses in a mouse model, and to demonstrate the clearance effects of this mineral against Porcine reproductive and respiratory syndrome virus (PRRSV) in experimentally infected pigs as an initial step towards the development of a feed supplement that would promote immune activity and help prevent diseases. In the mouse model, dietary supplementation with GB enhanced concanavalin A (ConA)-induced lymphocyte proliferation and increased the percentage of CD3+CD8+ T lymphocytes. In pigs experimentally infected with PRRSV, viral titers in lungs and lymphoid tissues from the GB-fed group were significantly decreased compared to those of the control group 12 days post-infection. Corresponding histopathological analyses demonstrated that GB-fed pigs displayed less severe pathological changes associated with PRRSV infection compared to the control group, indicating that GB promotes PRRSV clearance. These antiviral effects in pigs may be related to the ability of GB to increase CD3+CD8+ T lymphocyte production observed in the mice. Hence, this mineral may be an effective feed supplement for increasing immune activity and preventing disease.

Type II enteropathy-associated T-cell lymphoma: a clinicopathologic study / 中华病理学杂志

<p><b>OBJECTIVE</b>To study the clinicopathologic features, immunohistochemical findings, differential diagnosis and prognosis of type II enteropathy-associated T-cell lymphoma (EATL).</p><p><b>METHODS</b>Fourteen cases of type II EATL encountered in Department of Pathology, Nanjing General Hospital were retrospectively reviewed. The clinical data, histologic features, immunohistochemical findings and follow-up information were analyzed, with literature review.</p><p><b>RESULTS</b>There were altogether 12 males and 2 females. The median age of patient was 49 years. The sites of involvement included jejunum (10 cases) and ileum/colon (4 cases). The patients often presented with an abdominal mass, abdominal pain, diarrhea and constitutional symptoms such as fever, night sweating and cachexia. There was no clinical evidence of gluten-sensitive enteropathy. Histologically, the lymphoma cells showed full-thickness infiltration of the intestinal wall. They contained round hyperchromatic nuclei and pale cytoplasm. The stroma was minimally inflamed, with or without associated coagulative necrosis. A remarkable finding was the presence of villous atrophy, cryptal hyperplasia and intraepithelial lymphocytosis. Immunohistochemical study showed that the tumor cells expressed CD3, CD43 and CD8 (14/14). Some of them were also positive for CD56 (11/14) and CD30 (2/14). The staining for CD4, CD20, CD79a and myeloperoxidase was negative. A high proliferation index was demonstrated by Ki-67 immunostain. In-situ hybridization for EBER was negative. Follow-up data were available in 9 cases. The duration of follow-up ranged from 6 months to 36 months. Seven patients died within 14 months.</p><p><b>CONCLUSIONS</b>EATL is a rare type of lymphoma with intestinal involvement. Associated enteropathy is not demonstrated, in contrast to cases encountered in Nordic countries. A correct diagnosis requires evaluation of clinical manifestations, pathologic features and ancillary study results.</p>

Clinicopathologic features of intestinal natural killer/T-cell lymphoma / 中华病理学杂志

<p><b>OBJECTIVE</b>To study the clinicopathologic features, diagnosis and differential diagnosis of intestinal natural killer (NK)/T-cell lymphoma.</p><p><b>METHODS</b>The clinical features, histopathology, immunohistochemical findings and follow-up data of 14 cases of intestinal NK/T-cell lymphoma were retrospectively reviewed.</p><p><b>RESULTS</b>The male-to-female ratio was 9:5. The medium age of patients was 45 years. The sites of involvement included small intestine (6 cases), colon (6 cases) or both (2 cases). The main clinical manifestations were an abdominal mass, other gastrointestinal symptoms such as abdominal pain, as well as systemic symptoms such as fever and cachexia. Intestinal perforation complicated by acute peritonitis might occur in advanced disease. Histologically, the intestinal wall showed full-thickness infiltration by medium-sized atypical lymphoid cells with pleomorphic nuclei, prominent inflammatory background, angiocentric/angiodestructive growth pattern and coagulative necrosis. Immunohistochemical study showed that the tumor cells were positive for CD3ε, CD43, CD56, granzyme B and perforin. They were negative for CD20, CD79α and MPO. In-situ hybridization for Epstein-Barr virus encoded RNA (EBER) showed negative signals. A high proliferative index was demonstrated by Ki-67 immunostaining. Follow-up data of 8 cases were available, with duration of follow up ranging from 0.5 to 36 months. Five patients died within 20 months.</p><p><b>CONCLUSIONS</b>Extranodal NK/T-cell lymphoma, nasal-type primarily involving intestine is rare and tends to carry an aggressive clinical course. The relatively non-specific clinical manifestations of intestinal NK/T-cell lymphoma may result in misdiagnosis in some cases. A comprehensive evaluation of clinical manifestations, pathologic features and immunohistochemical findings is essential for definitive diagnosis.</p>

Influence of albumin and placement time outside the incubator on viability of isolated cytokine-induced killer cells: a simple method for detection / 南方医科大学学报

<p><b>OBJECTIVE</b>To study the influence of albumin supplementation in the medium and the placement time outside the incubator on the viability of isolated cytokine-induced killer (CIK) cells.</p><p><b>METHODS</b>CIK cells labeled with anti-CD3-FITC and anti-CD56-PE or with FQ-AE were observed under fluorescence microscope. The effect of albumin in the cell medium on the cell viability was analyzed using flow cytometry with Annexin V-FITC after different time lengths of placement.</p><p><b>RESULTS</b>The clones of CIK cells from the patient with esophageal carcinoma were small and scattered in the medium, but the clones from the patients with pancreatic cancer were large and densely distributed. CD3(+)CD56(+) cells could be detected under fluorescence microscope. The addition of albumin in the medium did not obviously affect cell apoptosis and death of CIK cells. CIK cells placed outside the incubator for less than 90 min showed a significant lower apoptosis rate than the cells placed for 150 min, whereas the cell death rate did not vary significantly with the placement time.</p><p><b>CONCLUSION</b>CIK cells from different cancer patients present with different growth pattern of the cells clones. Labeling with anti-CD3-FITC and anti-CD56-PE allows convenient counting of the newly generated CD3(+)CD56(+) CIK cells and FQ-AE labeling can be used for quantitative assessment of cell death. Albumin is not necessary in the medium of CIK cells. Prolonged placement (for over 90 min) of CIK cells outside the incubator should be avoided, and the placement time should be shorten as much as possible.</p>

Expression of PD-1/PD-L1 in peripheral blood mononuclear cells in lung cancer patients and its biological significance / 中华肿瘤杂志

<p><b>OBJECTIVE</b>To analyze the expression of co-stimulatory molecules PD-1/PD-L1 in peripheral blood mononuclear cells in lung cancer patients, and to explore its biological significance.</p><p><b>METHODS</b>One hundred and thirty-three lung cancer patients, 25 lung infection patients and 23 healthy donors were enrolled in this study. 100 µl of whole blood from these subjects were collected. Multi-color immunofluorescence staining and flow cytometry were used to detect PD-1/PD-L1 expression. The results were statistically analyzed.</p><p><b>RESULTS</b>The expression level of CD3⁺CD8⁺ T cells in the lung cancer patients was (38.83 ± 1.74)%, significantly lower than that in the control group [(43.25 ± 3.35)%, P < 0.05]. CD8⁺CD28⁺ T cell subset in the peripheral blood of lung cancer patients was (17.73 ± 1.21)% significantly lower than that of the healthy donors [(27.96 ± 2.72)%, P < 0.01]. The CD8⁺CD28⁻ T cell subset was (21.19 ± 1.92)% in the lung cancer patients, significantly higher than that of the healthy control group [(15.18 ± 2.93)%, P < 0.05]. The expression level of PD-1 on the surface of CD8⁺CD28⁺ T cells was (10.67 ± 1.12)% in the group of lung cancer patients, significantly higher than that of the control group [(5.32 ± 1.58)%, P < 0.01]. It was also found that the expression of PD-1 on CD8⁺CD28⁻ T cells was up-regulated in the group of lung cancer patients (7.46 ± 1.25)%, significantly higher than that of the healthy control group [(2.68+1.07)%, P < 0.01]. The expression level of PD-L1 on CD68⁺ cells in the lung cancer patients was (16.03 ± 2.06)%, significantly higher than that of the healthy control group [(9.32 ± 2.00)%, P < 0.05].</p><p><b>CONCLUSION</b>Up-regulation of PD-1/PD-L1 on peripheral blood cells in lung cancer patients negatively regulates the lymphocytes, inhibits the immune response for killing tumor cells, and promotes tumor development and immune escape.</p>