Autologous dendritic cells combined with cytokine-induced killer cells in the treatment of advanced non-small-cell lung carcinoma / 解放军医学杂志

Objective To evaluate the clinical efficacy and safety of dendritic cell (DC) combined with cytokine induced killer cell (CIK) in treatment of advanced non-small-cell lung carcinoma (NSCLC).Methods Peripheral blood mononuclear cells (PBMCs) were collected from 39 patients with advanced NSCLC,who were admitted to Affiliated Hospital of Academy of Military Medical Sciences,and cultured in vitro to produce DCs and CIK cells which,after phenotypic characterization by flow cytometry,were then returned to the patients.DCs were given subcutaneously on day 7,9,11 and 13 and CIK cells were given intravenously on day 11 and 13.The clinical efficacy and safety were analyzed before and after DCs-CIK cells treatment.Results Following up displayed that,in 39 patients with advanced NSCLC and eligible for evaluation,the objective response rate (ORR) was 30.8％ including 2 cases of completed response (CR) and 10 cases of partial response (PR),the disease control rate (DCR) was 69.2％.No significant difference existed pre-and post-treatment on the proportion of T cell subsets including CD3+CD4+CD8ˉ,CD3+CD4ˉ CD8+,CD3+CD19ˉ,CD3ˉCD 19+,CD3 CD16+CD56+,CD3+CD16+D56+,CD3+HLA-DRˉ,CD3+HLA-DR+ and CD3+CD28+CD8+ (P＞0.05),while obvious changes were found in Th1,Th2 and CD3+CD4+CD25+ T cells (Treg cells) (P＜0.05).No serious adverse events were observed.Conclusion Combined DCs-CIK cells immunotherapy provides a safe and effective treatment for patients with advanced NSCLC,improves the quality of life and relieves the probability of metastasis and recurrence.

Effects of body mass index and serum inflammatory cytokines on asthma control in children with asthma / 中国当代儿科杂志

<p><b>OBJECTIVE</b>To explore the effects of body mass index (BMI) and the levels of serum inflammatory cytokines on asthma control in children with asthma.</p><p><b>METHODS</b>One hundred and sixteen children with asthma were divided into three groups: normal (n=59), thin (n=31), and obesity (n=26) based on their BMI. The levels of interleukin-6 (IL-6) and tumor necrosis factor-alpha (TNF-α) were determined using ELISA, and the level of high-sensitivity C-reactive protein (hs-CRP) was measured by immunoturbidimetric assays. Asthma control status in each group was evaluated by the Childhood Asthma Control Test (C-ACT) after 4 weeks of treatment.</p><p><b>RESULTS</b>The serum levels of IL-6, hs-CRP, and TNF-α were highest in the obesity group, followed by the thin group and the normal group (P<0.05), while the C-ACT score was highest in the normal group, followed by the thin group and obesity group (P<0.05). The normal group had significantly higher complete controlled and partially controlled rates than the thin and obesity groups (P<0.05); however, there were no significant differences between the thin and obesity groups (P>0.05). The levels of IL-6, hs-CRP, and TNF-α were negatively correlated with the C-ACT score (P<0.05). There were no significant correlations of BMI with the C-ACT score and levels of IL-6, hs-CRP, and TNF-α (P>0.05).</p><p><b>CONCLUSIONS</b>When BMI is too high or too low, the levels of serum inflammatory cytokines are all increased, which is harmful to asthma control. Maintaining a healthy weight in children with asthma may reduce the levels of serum inflammatory cytokines and improve the asthma control rate.</p>

Expression of NOV and BNIP3 gene in mouse myelomonocytic leukemia and its significance / 中国实验血液学杂志

This study was aimed to investigate the expression level of NOV and BNIP3 mRNA in mice myelomonocytic leukemia (AML-M(4)) and its significance. The mice were inoculated intravenously with myelomonocytic leukemia cells of WEHI-3, and divided randomly into chemotherapy group and control (untreated) group. Bone marrow samples were then collected from both groups at different times. The NOV and BNIP3 mRNA expression were detected by TaqMan quantitative real-time reverse transcription polymerase chain reaction (qRT-PCR), and the relationship between these expression levels and clinical significance in leukemia incidence and progression were analyzed with β-actin as the housekeeping gene. The results showed that the mean values of NOV and BNIP3 increased gradually from 2 weeks after inoculation and achieved highest level at death in control group. Expression level of NOV increased from 1.85E-05 before inoculation to 3.57E-02 at death (p < 0.05), and BNIP3 from 3.44E-03 to 3.48E-02. While 2 gene expression in the chemotherapy group decreased quickly to 2.51E-05 and 1.58E-03 (p < 0.05) respectively after chemotherapy, which were close to the level before inoculation (p > 0.05). The 2 gene expressions again rose at relapse, and difference of expression level between 2 group at death were no statistically significant (p > 0.05). It is concluded that the expression of NOV and BNIP3 in leukemia AML-M(4) is significantly higher than that in normal controls, of which high level expression is an important factor in the development of leukemia. Close relation between the therapeutic effect and expression level of these two genes suggests the great value in prognostic evaluation and MRD detection.

Relationship between WT1-specific T-cell subsets and graft-versus-host disease after nonmyeloablative allogeneic transplantation / 中华血液学杂志

<p><b>OBJECTIVE</b>To explore the relationship between WT1-induced T-cell subsets and graft-versus-host disease (GVHD) after nonmyeloablative allogeneic hematopoietic stem cell transplantation (NST).</p><p><b>METHODS</b>Peripheral blood mononucleated cells (PBMCs) from 19 patients who expressed WT1 and developed GVHD after NST were simulated by WT1126-134 peptide in vitro, and proportions of WT1-induced-T-cell subsets (Tc1, Tc2, Th1, Th2 cells) before and after transplant were detected by intracellular cytokine staining (ICCS) assay. WT1-specific CD8(+) CTLs of 14 patients with HLA-A*0201 were detected by HLA-A*0201/WT1 pentamer.</p><p><b>RESULTS</b>(1) 17 of 19 patients developed GVHD, among whom proportions of Tc1 and Th1 cells, achieved peak value in 16 patients at occurrence of GVHD (P = 0.039); (2) The peak proportions of Tc1 and Th1 cells in patients with aGVHD above grade II were higher than those with grade I, but being no statistical difference (P = 0.900 and P = 0.140, respectively); (3) The peak proportion of Th1 cells (P = 0.004), but not Tc1 cells (P = 0.060) in patients with extensive cGVHD was significantly higher than that in patients with limited one; (4) Proportions of Tc1, Th1 and WT1(+)CD8(+)CTL in patients without GVHD were similar to those in patients with Grade I aGVHD, but lower than those in aGVHD above grade II.</p><p><b>CONCLUSION</b>GVHD promotes the generation of WT1-induced GVL effect, and the intensity of the latter maybe correlated with the intensity of GVHD, especially cGVHD. Th1 cells play a more important role in the enhancement of WT1-induced GVL effect in extensive cGVHD patient than in limited cGVHD patients.</p>

Application of HLA-A*0201/WT1 pentamer combined with intracellular IFNgamma+ staining in detecting circulating WT1 specific T cells in leukemia / 中国实验血液学杂志

This study was purposed to investigate the value of combination of pentamer and intracellular IFNgamma staining in the qualitative and quantitative detection of circulating antigen-specific T cells. WT1 expressions in 14 HLA-A*0201+ patients and their matched donors were detected by RT-PCR, and circulating WT1 specific T cells were assayed by HLA-A*0201/WT1 pentamer combined with intracellular IFNgamma+ staining. The results showed that the low level of WT1 expression was found only in 2 cases out of 14 donors, but different levels of WT1 expression could be observed in all leukemic patients. The WT1+CD8+ CTL and WT1+IFNgamma+ cells did not detected in all 14 donors, but WT1+CD8+ CTL cells in 2 patients and WT1+IFNgamma+ cells in 3 patients could be detected before transplantation respectively, there was no significant difference between them, while the WT1+CD8+ CTL cells and WT1+IFNgamma+ cells both could be detected in all 14 patients after transplantation, the positive detection rate after transplantation was obviously higher than that before transplantation. The WT1+CD8+ and WT1+ IFNgamma+ cells could be detected within 30 days after transplantation, but the positive detection rate of WT1+IFNgamma+ cells was higher than that of WT1+CD8+ CTL cells (p=0.014). The median peak value of WT1+CD8+ CTL cells was 0.18% in 14 patients, and the median peak value of WT1+IFNgamma+ cells was 0.83% in 14 patients, the later was significantly higher than former. The median peak time of WT1+CD8+ CTL cells was 75 days after transplantation, while the WT1+IFNgamma+ cells was 105 days after transplantation, there was no significant difference between them. It is concluded that pentamer and intracellular IFNgamma staining may effectively detect circulating WT1 specific T cells in leukemic patients, and the combination of these two methods profit to the exact qualitation and quantitation of circulating antigen-specific T cells.

Establishment and evaluation of quantitative analysis method for donor chimerism of mice / 中国实验血液学杂志

This study was aimed to explore the effectiveness of sequential and quantitative detection method for analysing donor chimerism (DC). In order to simulate a mouse model of haploidentical stem cell transplantation, C57BL/6 male mice were used as donors, while CB6F1 female mice were used as recipients and were divided into 2 groups. The two groups of recipients were irradiated with 2 Gy and 6 Gy from (⁶⁰CO gamma-ray source respectively, and then were inoculated intravenously with bone marrow cells (BMCs) and spleen mononuclear cells (SPMNCs). Quantitative analyses of DC were performed with real-time PCR or flow cytometry (FCM) on different days after transplantation. The results showed that real-time PCR and FCM both have advantages and disadvantages in the detection of DC. When DC amount in group of 6 Gy was > 90% with stable macrochimerism for more than 3 months, the efficacy of detection by FCM was well; while the DC amount in group of 2 Gy was below 10% and gradually transformed to different forms of microchimerism until disappeared, in which condition the use of real time-PCR was more appropriate. It is concluded that FCM can detect macrochimerism with high accuracy but would fail when DC amount is less than 1% due to sensitivity limitation, while the real time-PCR is more sensitive for detecting microchimerism but lack of accuracy for detecting macrochimerism. Combination of the two methods can afford sensitive and accurate tool for quantitative analysis of chimerism in mouse model of transplantation and adoptive immunotherapy.

Changes of lymphocyte subsets in acute leukemia patients after HLA-mismatched nonmyeloablative hematopoietic stem cell transplantation / 中国实验血液学杂志

This study was purposed to investigate the reconstitution of immune system in patients with acute lymphocyte leukemia (ALL) or acute myeloid leukemia (AML) after HLA-mismatched nonmyeloablative hematopoietic stem cell transplantation (NHSCT) and its relation with infection and GVHD. 6 ALL and 4 AML patients having HLA-mismatched related donors received the nonmyeloablative precondition regimen composed of fludarabine (Fln), ATG, Ara-C, CTX and total body irradiation (TBI) in dose 2 Gy. The GVHD was prevented and treated by CsA, anti-CD25 antibody and mycophenolic mofetil (MMF) before and after transplantation. The flow cytometry was used to detect the changes of total T cells, help/inducer T cells, suppressor/killer T cells, gamma/delta T cells, B cells, NK cells, NKT cells, regulatory T cells, activated T cells, naive T cells, memory T cells and ratio of CD4/CD8 in patients with remission resulting from chemotherapy before transplantation, and analyse the relation of immunofunctional cells to infection and GVHD after transplantation, compare the difference in recovery of immune system of ALL and AML patients. The results showed that the recovery of total lymphocytes and lymphocyte subsets displayed one's own regular pattern. As compared with patients without GVHD, the counts of lymphocyte subsets in patients with GVHD was higher, while the counts of gamma/delta T cells, regulatory T cells, NK cells, the counts of B cells, NK cells, naive cells and CD4/CD8 ratio as well as the counts of B cells, naive T cells and NK cells were lower at 1 month, 2 - 3 months and 6 - 8 months after transplantation respectively. The total T cells and subsets recovered slowly, but NK cells and NKT cells recovered rapidly in patients with infection at early period after transplantation, the B cells and naive B cells recovered rapidly at 3 months after transplantation. There was no difference in lymphocyte recovery between ALL and AML patients. It is concluded that the analysis of each lymphocyte subsets may indirectly show the recovery of thymus function in patients, the changes of NK cells, B cells and naive T cells have an important significance for identifying and forecasting the GVHD and infection.

Acute graft versus host disease in non-myeloablative allogeneic stem cell transplantation / 中国实验血液学杂志

The objective of this study was to explore the occurrence and clinical features of acute graft versus host disease (aGVHD) in non-myeloablative stem cell transplantation (NAST). 19 cases developed aGVHD out of 71 cases with NAST in recent years were analyzed retrospectively. Out of 19 cases, 9 males and 10 females at the median age of 38 (18-59), 16 cases with grade I-II aGVHD, 3 cases with grade III-IV aGVHD. The results indicated that the incidence of aGVHD in NAST was 26.7% (19/71), and severe aGVHD was 4.2%, the median onset time was 58 days (17-240 days) after transplantation. Skin and especially the intestine were the main target organs of aGVHD, while diarrhea occurred as the first symptom in 7 cases, 3 cases showed mixed acute and chronic GVHD involving more locations at the same time. aGVHD occurrence was 38.2% in those patients with full donor chimerism (FDC) and 16% in patients with the mixed chimerism (MC). It is concluded that aGVHD in NAST is less in occurrence, lighter in severity and later in time, but higher occurrence in those with early FDC, which intestine and skin are the main target organs. The clinical course is prolonged and easily complicated with severe infection in the later phase. Early combined therapy with powerful supportive treatment is necessary.

Comparison of efficiencies mobilizing stem cells into peripheral blood in healthy donors by different schemes with G-CSF / 中国实验血液学杂志

The aim of study was to select the best scheme of G-CSF to mobilize peripheral stem/progenitor cells in healthy donors. The clinical data of 60 cases received nonmyeloablative allogenic hematopoietic stem cell transplantation was analyzed retrospectively. The results indicated that the counts of MNC and CD34(+) cells were significantly higher in the 10 microg/(kg.d) group than that in the 5 microg/(kg.d) group (P < 0.05). The counts of MNC and CD34(+) cells which were collected after day 4 or 5 in the 10 microg/(kg.d) groups were not significantly different. The percentage of CD3(+) cells, CD4(+) cells and CD8(+) cells were not different in different groups. It is concluded that the scheme using 10 microg/(kg.d) G-CSF is more efficient than that in the 5 microg/(kg.d) group in mobilizing stem cells. It may reduce days for mobilization and decrease expense for collection of cells after 4 days of mobilization. Scheme using 10 microg/(kg.d) G-CSF for cells collecting after 4 days is more efficient.

ABO-incompatible nonmyeloablative allogeneic peripheral blood stem cell transplantation / 中国实验血液学杂志

To explore the effects of ABO incompatibility between recipient and donor on HLA-matched nonmyeloablative allogeneic peripheral blood stem cell transplantation (NAST), a retrospective, cohort study was performed. Among 24 HLA-matched NAST, 15 were major ABO-incompatible and 9 minor. Control group included 24 HLA-matched NAST with ABO-compatible grafts. Nonmyeloablative conditioning regimens consisted of CTX, Ara-C and ATG. The patients were given cyclosporine A and mycophenolate mofetile for prophylaxis of acute GVHD. The ABO-incompatible patients received grafts depleted erythrocytes by hydroxyethyl starch (HES) sedimentation. The results showed that successful and stable engraftment was established in 23 patients. No recipient developed clinically immediate hemolysis during graft infusion, but 2 recipients experienced delayed hemolysis attributable to the ABO incompatibility. The median time of granulocyte counts >0.5 x 10(9)/L and platelet >30 x 10(9)/L was 11 and 14.9 days, respectively. In ABO major incompatible group, the onset of erythropoiesis after NAST was delayed. One out of 10 recipients with blood group "O" in this group developed pure red cell aplasia (PRCA), lasting 5 months. The acute GVHD occurred in 7 out of the 24 patients. The chronic GVHD occurred in 5 of 21 cases. Relapse was observed in 2 patients with acute leukemia. The actuarial probability of disease-free survival at 2 years was 63.3%. In conclusion, ABO-incompatible grafts for NAST have no adverse effect on engraftment, recovery of platelets, incidence of GVHD, relapse rate or survival. ABO-incompatible NAST is fairly safe if there is indication, however, the onset of erythropoiesis is delayed when major ABO mismatched.

Diagnosis and treatment of invasive pulmonary aspergillosis in malignant hematological diseases-3 cases reported / 中国实验血液学杂志

Invasive pulmonary aspergillosis is difficult to diagnose and a critical ill with high mortality. In this paper, the diagnosis and treatment of invasive pulmonary aspergillosis complicated in 3 cases of hematological malignancy (2 acute leukemias and 1 MDS-RA) were retrospectively analysed. All patients had histories of hypoimmunity and were received prophylactic antifungal treatment. Pulmonary aspergillosis infection still occurred and confirmedly diagnosed by sputum examination. After 7 to 14 days of combination treatment of liposomal amphotericin B, itraconazole and flucytosine, 2 cases were cured and another showed effective. In conclusion, early diagnosis and treatment of invasive pulmonary aspergillosis are very critical and the therapeutic effectiveness of combined scheme with liposomal amphotericin B, itraconazole and flucytosine is very effective for pulmonary aspergillosis.

Comparison of conditioning regimens containing or no fludarabine in nonmyeloablative allogeneic peripheral blood stem cell transplantation / 中华血液学杂志

<p><b>OBJECTIVE</b>To investigate the therapeutic effect of conditioning regimen containing fludarabine in nonmyeloablative allogeneic peripheral blood stem cells transplantation (NAST) in the treatment of hematological diseases.</p><p><b>METHODS</b>Thirty-six patients with acute leukaemia, severe aplastic anaemia, MDS and myelofibrosis received NAST from HLA matched donors' G-CSF mobilized peripheral blood stem cells after nonmyeloabalative conditioning. The conditioning regimen consisted of CTX, Ara-C, CsA, anti-CD(3) antibody or anti-thymocyte globulin and with or without fludarabine. GVHD prophylaxis was performed with cyclosporine combined methotrexate (no MMF group, n = 5) or mycophenolate mofetil (MMF group, n = 31).</p><p><b>RESULTS</b>All of the treatment was generally well tolerated and all cases achieved engrafted of the donor cells. In fludarabine group, engraftment was observed in 87.5% (14/16) patients with complete donor chimerism, graft failure was 12.5% (2/16) and in no fludarabine group, 80% (16/20) and 20% (4/20), respectively. The incidence of acute GVHD (grade I - IV) was 27.8% (10/36) and chronic GVHD 22.2% (8/36). In fludarabine group, grade I - II aGVHD was 37.5%, in no fludarabine group, 20%. cGVHD was 12.5% in fludarabine group and in no fludarabine group 30%, respectively. Interstitial pneumonia (IP) was observed in 16.7% (6/36) of the patients, being 18.7% (3/16) and 15% (3/20) in fludarabine and no fludarabine group, respectively. Overall survival rate was 80.5% (29/36) with a median follow-up of 13 months.</p><p><b>CONCLUSIONS</b>There was no significant difference between fludarabine based (n = 16) and non-fludarabine based conditioning regimen (n = 20) in NAST for the treatment of hematological diseases, regarding for incidence of GVHD, IP, engraftment and survival.</p>

The clinical research of nonmyeloablative allogeneic peripheral blood hematopoietic stem cells transplantation for hematological diseases / 中华血液学杂志

<p><b>OBJECTIVE</b>To explore the significance of nonmyeloablative allogeneic peripheral blood hematopoietic stem cell transplantation in the treatment of hematological diseases.</p><p><b>METHODS</b>A nonmyeloablative conditioning regimen consisted of CD(3) monoclonal antibody, cyclosporine A, cyclophosphamide and cytarabine was used for allogeneic stem cell transplantation in 33 patients with hematological diseases. Of them, 11 were acute leukemia (AL) in first complete remission (CR(1)), 4 AL-CR(2) approximately 3, 3 refractory AL, 4 severe aplastic anemia (SAA), 7 chronic myeloid leukemia (CML), 2 myelodysplastic syndrome, 1 each of chronic lymphocytic leukemia (CLL) and myelofibrosis.</p><p><b>RESULTS</b>All 33 patients passed the hematopoietic suppression stage smoothly and achieved engraftment of the donor cells. There were 24 cases of full donor chimerism (13 cases converted from mixed chimerism), 4 mixed chimerism (MC) and 5 developed graft rejection. Of the 33 cases, 7 (21.2%) developed acute GVHD and chronic GVHD, 25 (75.8%) still live and 8 (24.2%) died.</p><p><b>CONCLUSIONS</b>Nonmyeloablative allogeneic peripheral blood stem cells transplantation is a safe, less toxic and curative approach for patients with hematological disease.</p>

Nonmyeloablative Allogeneic Peripheral Blood Hematopoietic Stem Cell Transplantation for Therapy of Hematologic Diseases / 中国实验血液学杂志

The objectve of this investigation was to explore the use of nonmyeloablative allogeneic peripheral blood stem cell tranplantation for treatment of hematologic diseases. Six patients were included: 3 cases with acute leukemia in first complete remission (2 AML and 1 ALL), 2 severe aplastic anemias and 1 chronic myeloid leukemia in chronic phase. All of the 6 cases were received HLA-identical siblings donor peripheral blood stem cell transplantation after a nonmyeloablative conditioning. The donor cells were engrafted in all patients (3 cases were full engrafment of donor cells and 3 were mixed chimerism). Hematopoietic recovery was appeared in all of the cases (ANC recovered to more than 0.5 x 10(9)/L and platelet count to more than 30 x 10(9)/L on day 9 to day 21 and day 14 to day 28 after transplantation, respectively). Two patients developed III or IV degree GVHD. Our preliminary results suggest that the procedure is more safe, efficient and less complications than myeloablative conditioning regimens and represents another new approach in the management of patients with hematologic diseases.