Several hot questions such as selection of patients and favorable time of transplantation, design of conditioning regimen, monitoring of chimerism and complication of graft versus host disease (GVHD) in allogeneic hematopoietic stem cell transplantation following reduced intensity regimen (RIC-HSCT) as salvage therapy for refractory leukemia are discussed in this article. More and more investigators began to recognize that it was not a fundamental to undergo transplantation in complete remission for patients with refractory leukemia, since it may expend patients' physical status. RIC-HSCT may be substantially considered as a kind of adoptive immunotherapy, so that immunologic attacking targets and the latency of immunoreactian must be considered when making the decision to use, lacking of attacking targets and rapidly growing diseases seemed to be less susceptible to control. Commonly used reduced intensity regimens differed significantly, but it was clear that dose intensity was very important in refractory leukemia. In fact, many investigators used intermediate dosage between criteria of non-myeloablative and conventional myeloablative regimens. Complete donor chimerism is the hallmark of engraftment but often delayed in RIC-HSCT. Since sustained complete donor chimerism induced persistent graft-versus-leukemia (GVL) effect play an important role in patients' long-term survival, it was recommended that sensitive techniques (eg. STR-PCR) should be used to analysis chimerism and should be measured more frequently (every 2-4 weeks), and lineage-specific chimerism (eg. T cell) analysis was also recommended. As compared with traditional HSCT, the incidences of acute and chronic GVHD are similar and the onset of GVHD is associated with the GVL effect. Decrease or interruption of immunosuppressive drugs early after transplantation and donor lymphocyte infusion may facilitate transformation to complete donor chimerism, so that it may benefit patients with advanced disease at time of transplantation from avoiding disease relapse in one hand, but may induce GVHD in the other hand.

Objective To investigate the role of micro-RNAs (miR-101 and miR-125a-5p) in autophagy of lipopolysaccharide (LPS) derived human THP-1 macrophages.Methods Human monocytic leukemia cell line THP-1 was cultured in vitro,and it was differentiated into macrophages after being induced with phorbol (50 μg/L) for 48 hours.THP-1 macrophages were stimulated with LPS in 0,250,500,1 000 μg/L respectively for 12 hours,miR-mimic was transfected into THP-1 macrophages as induced by Lipofectamine RNAiMAX,and the transfection efficiency of miRNA was determined with fluorescence microscopy.Enzyme linked immunosorbent assay (ELISA) was used to determine the levels of tumor necrosis factor-α (TNF-α) and monocyte chemotaxis protein-1 (MCP-1) in the supernatants of culture.Western Blot was used to detect the protein expressions of autophagy proteins ATG4D,Beclin1,and LC3 Ⅱ.The expression levels of miR-101 and miR-125a-5p were determined by quantitative reverse transcription-quantitative polymerase chain reaction (RT-qPCR).Results ① The releasing levels of TNF-α and MCP-1 induced by LPS with 250,500,1 000 μg/L were significantly increased as compared the cells without LPS stimulation [TNF-α (ng/L):1 336.1 ± 18.5,1 340.6±24.8,1 364.5± 14.9 vs.47.6±4.4;MCP-1 (ng/L):996.3 ±51.3,934.6±84.3,974.2±69.5 vs.21.3±6.5,all P ＜ 0.01],but no significant differences were found among the three LPS stimulation groups.The protein expressions of ATG4D,Beclin1 and LC3 1Ⅱ were up-regulated in the presence of different LPS concentrations (0,250,500,1 000 μg/L) for 12 hours in THP-1 macrophages (when compared with the cells without LPS stimulation,t value of ATG4D was 8.103,38.410,52.020,P value was 0.015,0.001,＜ 0.001;t value of Beclin1 was 3.026,5.328,3.482,P value was 0.047,0.034,0.037;t value of LC3 Ⅱ was 3.836,6.200,4.665,P value was 0.018,0.003,0.010),and the optimal concentration was 500 tg/L LPS.When THP-1 macrophages were stimulated with 500 μg/L LPS for 12 hours,the expression levels of miR-101 and miR-125a-5p were down-regulated significantly as compared with the cells without LPS stimulation [miR-101 (2-△ △Ct):0.68 ± 0.08 vs.1.95 ±0.26,t =8.047,P =0.001;miR-125a-5p (2-△ △Ct):0.23 ± 0.06 vs.1.69± 0.42,t =5.975,P =0.004].② The higher transfection efficiency was showed under fluorescence microscope.Westem Blot results showed the protein expressions of ATG4D,Beclin1 and LC3 Ⅱ were down-regulated as induced by an over-expression of miR-101 or miR-125a-5p in THP-1 macrophages,and more obviously down regulated by co-transfected with miR-101 and miR-125a-5p (compared with negative control group,t value was 14.550,5.855,14.180,P value was 0.005,0.014,＜ 0.001).Conclusion miR-101 and miR-125a-5p can inhibit the autophagy in LPS challenged THP-1 macrophages,and the potential mechanism might be related to target regulation of ATG4D.

Background and purpose:The patients with aggressive lymphoma who have a poor prognosis and unlikely to be cured with conventional chemotherapy. This study was aimed to evaluate the effect of high-dose etoposide in mobilization followed auto-SCT in treating refractory lymphoma. Methods:40 patients [median age 33 (13-61) years] with refractory non-Hodgkin’s lymphoma (NHL, n=32) or Hodgkin’s lymphoma (HD, n=8) received high-dose etoposide [VP16 10-15 mg/(kg·d)×2 d] in mobilization in our center. Remission status prior to mobilization was PD (n=40). The use of such granulocyte colony-stimulating factor [G-CSF, 5-10μg/(kg·d)] mobilized peripheral blood stem cells (PBSC) after high-dose etoposide until the end of leukapheresis. Peripheral blood stem cell was collected and frozen in-80℃refrigerator. All these patients received auto peripheral blood stem cell transplantation (auto-PBSCT). Conditioning regimen was BEAM (n=19, 47.5%) or CBV (n=21, 52.5%). Results:Twenty-eight pa-tients (70%) were assessable for response after high-dose etoposide at a median pretreatment time of 39 days (range 17-172 days), 12 patients (30%) had no response. Median follow-up of 28 (4-66) months, 16 patients (40%) reached CR after auto-PBSCT. Fifteen of the 28 patients (53.6%) who had response to high-dose etoposide reached CR, 4 patients (14.3%) reached PR, 9 patients (32.1%) succumb to progression of disease. One of the 12 patients (8.3%) who had no response to high-dose etoposide reached CR, 1 patients (8.3%) reached PR, 10 patients (83.4%) succumb to progression of disease. The estimated 1-year OS and EFS were 69%and 56.7%respectively, 2-years OS and EFS were 63%and 52%respectively. The prognosis of the patients who had no response to etoposide was poor. The estimated 1-year OS and EFS were 25%and 16.7%respectively. Two group of comparison differences have statistics signiifcance (P<0.01). Conclusion: High-dose etoposide could be used in refractory lymphoma as rescue therapy in mobilization. It can increase the EFS and OS of patients who had response. The hematopoietic stem cells collection and hematopoietic reconstitution are not affected by etoposide.

Objective To retrospectively analyze the effect of preemptive treatment on cytomegaloviras (CMV) infection in patients receiving allogeneic hematopoietic stem cell transplantation (allo-HSCT). Methods The data of one hundred and three patients who underwent alIo-HSCT with preemptive treatment to prevent CMV associated diseases were retrospectively analyzed. Fluorescence quantitative PCR was used to detect CMV-DNA. The incidences of CMV viremia and CMV associated diseases were analyzed. Results CMV viremia was confirmed 63 times in 51 of the 103 patients. The incidence of CMV viremia was 49. 5% and the median time of onset was 40 days after transplantation. All the patients with CMV viremia received preemptive antiviral therapy and 19 of them developed CMV associated diseases, including 14 hemorrhagic cystitis, 3 CMV associated pneumonia and 2 CMV associated enteritis. The total incidence of CMV associated diseases was 18. 4%. After treatment with ganciclovir and/or foscarnet, 60 of the 63 times of CMV viremia disappeared. One patient was not included in the analysis because he died of intracranial hemorrhage and GVHD only 3 days after the treatment. The total response rate was 96. 8% (60/62). The remaining two cases who did not respond to treatment died of CMV associated pneumonia in combination with acute GVHD. The direct mortality rate of CMV infection was 1.9% (2/103). Conclusion The incidences of CMV viremia and CMV associated diseases do not increase in patients receiving preemptive therapy as compared with those receiving prophylaxis therapy. Preemptive treatment can not only prevent the progression of CMV viremia to CMV associated diseases in majority of the cases but also control CMV associated diseases effectively.

Objective To evaluate the value of the new ovarian function marker serum anti-Mullerian hormone (AMH)and to clarify the effect of hysterectomy on the ovarian function of the younger women.Methods The serum AMH,follicle stimulating hormone (FSH)and luteinizing hormone (LH)levels in 35 women suffered uterus benign lesion aged 36-45 years and 35 women suffered hysteromyoma without operation were measured at different time.And the ovarian arterial blood flow resistance index (RI ) was measured by Doppler ultrasound. Results Compred with before operation, the serum AMH levels of the patients in hysterectomy group 2 d and 3 months after operation were significantly decreased (P<0.05 or P<0.01).Compared with control group,the serum AMH levels of the patients in hysterectomy group 2 d and 3 months after operation were also decreased (P<0.05).The ovarian arterial blood flow RI of the patients in hysterectomy group 1 and 3 months after operation were increased compared with control group (P<0.05).There were no significant differences in the serum FSH and LH levels in each group between different time points (P>0.05).Conclusion Hysterectomy can affect the ovarian function,and the serum AMH level 3 months after operation is decreased and the ovarian arterial blood flow RI is increased.AMH is superior to FSH or LH in evaluating the changes of ovarian function.

Objective To evaluate the response rate and survival rates of refractory or relapsed Hodgkin lymphoma (HL) and grey zone lymphoma patients treated with autologous peripheral blood stem cell transplantation (APBSCT).Methods From January 2004 to August 2012,30 HL and grey zone lymphoma patients were retrospectively analyzed.Statistical analysis was done to explore the long term outcome and prognostic factors of patients treated with APBSCT.Among all patients,the median age at transplantion was 30 (13-55) years old.Patients were major with nodular sclerosis HL and in stage Ⅲ/Ⅳ.Results Every patient had a successful collection.The median MNC cell dose infused was 6.8×108/kg [range (1.0-13.8)×108/kg] and median CD34+ cell dose infused was 6.3×106/kg [range (0.6-20.6)×106/kg].Median time to neutrophil engraftment was 9 days (range 8-12 days).28 patients were evaluable after transplantation with a median follow-up of 18.5 months (range 2.5-95.0 months).The overall response rate was 89.3 ％ [CR 64.3 ％ (18/28),PR 25.0 ％ (7/28)].The overall survival (OS) rate and progression free survival (PFS) rate at 5 year would be 78 ％ and 58 ％ for all patients.3 in 7 patients with no remission after salvage chemotherapy with rituximab plus chemotherapy before APBSCT got CR and 2 got PR.Univariate analysis showed that disease status and the number of replacement types of chemotherapy prior to transplantation affected OS,the history of radiotherapy prior to transplantation affected PFS.Conclusion APBSCT can increase CR rate,prolong survival time in patients with refractory or relapsed HL and grey zone lymphoma.Rituximab plus chemotherapy as a salvage therapy could raise CR rate before APBSCT.Chemosensitivity before transplantation affect outcome with APBSCT.Changing many types of chemotherapy is adverse for APBSCT.Salvage radiotherapy before APBSCT is not recommended.

Objective To evaluate the efficacy of anti-CD20 monoclonal antibody (Rituximab) combined with autologous hematopoietic stem cell transplant (ASCT) in treatment of the patients with B cell non-Hodgkin lymphoma (NHL). Methods Twenty-one patients with B-cell NHL(CD20 positive) received ASCT with Rituximab at the dose of 385 mg·m-2·d-1 on day 1 and day 8 of mobilization,and day -1 and day +7 of conditioning regimen. Among the 21 patients receiving chemotherapy before the transplant, five cases achieved complete response (CR), eleven cases achieved partial remission (PR), and 5 cases had the progression of disease (PD) after many cycles of chemotherapy. Results The median follow-up was 24 months (1-68 months) in the present study. No relapse occurred among the 5 patients in CR before the transplant. Only one of the 11 PR patients relapsed 6 months post-transplantation. Three of the 5 PD patients died. Four of 21 cases (19 %) were documented as recurrence and death, the other 17 cases remained alive and disease-free. Both 2-year EFS and OS of these cases were 81%. No harmful effect of Rituximab was observed on the quality and quantity of collected stem cells as well as hematopoietic recovery post SCT. Conclusion The efficacy of ASCT with Rituximab in vivo purging in the patients with B-cell NHL was determined mainly by the disease status before transplant. The approach may be used as consolidation therapy to achieve long-term survival and increase the curable rate for patients in CR before transplant, and as intensification therapy to increase the remission rate and prolong the EFS and OS of the patients in PR. Rituximab did not show any adverse effect on collection and reconstitution of hematopoietic stem cells.

Objective The aim of this prospective study was to evaluate the changes in the ovarian reverse after myomectomy based on serum anti-Mullerian Hormone (AMH) levels. Methods This is a prospective longitudinal observational study. Serum AMH levels were measured at the baseline and 2 day , 3months after myomectomy in 35 women aging from 36 to 45years.Follicle stimulate hormone(FSH) and luteal hormone(LH) were measured at the same time. 35 women of the same age with fibroid who did not undergo operation were selected as control group. Result (1)AMH level is (1.54 ± 0.95)ng/mL,(1.18 ± 0.77)ng/mL,(1.50 ± 0.58 )ng/mL at 0 day, 2 days and 3 months after operation. AMH level decreased significantly at 2 days after operation (P < 0.05) and increased gradually 3 months after operation, but showed no significant change (P > 0.05).(2) Significant differences in the serial change of AMH levels existed at each time point between myomectomy group and control group (P <0.05). No significant differences in FSH or LH levels existed at each time point. Conclusion AMH is may be superior to FSH or LH in evaluating the changing of ovarian reverse. The study suggests that myomectomy affect the ovarian function for up to 3 months post-operatively , and hemorrhage during and after operation may decrease serum AMH levels.

Objective To examine the distribution of bacterial species and antimicrobial susceptibility profile of pathogens in febrile neutropenic patients.Methods A total of 355 bacterial strains were isolated from febrile neutropenic patients in Shanghai General Hospital from January 2005 to December 2012. Antimicrobial susceptibility testing was done by Kirby-Bauer method. The susceptibility testing results were analyzed according to CLSI 2014 breakpoints.Results Gram-negative bacteria accounted for 70.4% of the 355 isolates, while gram-positive organisms accounted for 29.6%. The most common bacterial species werePseudomonas aeruginosa, Klebsiella pneumoniae, Acinetobacter baumannii, Escherichia coli, Stenotrophomonas maltophiliaand Staphylococcus haemolyticus. Non-fermentative bacteria accounted for 53.2% of all the gram-negative bacterial isolates. All theEnterococcus and
Staphylococcus isolates were susceptible to linezolid, vancomycin and teicoplanin. All theStaphylococcus strains were resistant to methicillin.P. aeruginosa isolates were relatively more susceptible to cefoperazone-sulbactam, piperacillin-tazobactam and cefepime (>70%) than imipenem (40.8%) and meropenem (59.2%). All theK. pneumoniae isolates were susceptible to imipenem and meropenem and more than 70% of the isolates were susceptible to cefoperazone-sulbactam, amikacin. More than 80% of theA. baumannii isolates were susceptible to carbapenems, cefoperazone-sulbactam, amikacin, ciprolfoxacin and aminoglycosides. All the E. coli isolates were susceptible to carbapenems and more than 70% were susceptible to cefoperazone-sulbactam and ceftazidime. More than 90% of theS. maltophilia strains were sensitive to levolfoxacin, minocycline, cefoperazone-sulbactam and trimethoprim-sulfamethoxazole.Conclusions Our data suggest that gram-negative bacteria, especiallyEnterobacteriaceae and non-fermentative bacteria, are still the primary pathogens in febrile neutropenic patients. Antimicrobial resistant strains are prevalent. Such data of bacterial species and antimicrobial susceptibility proifle of pathogens in febrile neutropenic patients are useful for empirical antimicrobial therapy of such infections.

Objective To evaluate the efficacy of haploidentical allogeneic hematopoietic stem cell transplantation (Haplo-HSCT) combined with third-party umbilical cord blood (UCB) infusion in treatment of high-risk lymphoblastic malignancies. Methods The clinical data of 20 patients with high-risk lymphoblastic malignancies who received Haplo-HSCT from April 2012 to April 2015 in Shanghai General Hospital were retrospectively analyzed, which were compared with the data from 15 patients who underwent matched unrelated donor HSCT (MUD-HSCT) or 14 matched sibling donor HSCT (MSD-HSCT) during the same period. The efficacy of Haplo-HSCT combined with UCB infusion in treatment of high-risk lymphoblastic malignancies was evaluated. The preparative regimen mainly consisted of teniposide, cyclophosphamide and total body irradiation (TBI). Graft versus host disease (GVHD) preparative regimen included cyclosporine and a short term of methotrexate. The patients who received Haplo-HSCT combined with UCB infusion and MUD-HSCT were treated with antithymocyte globulin (ATG). Results After the transplantation, one patient in MUD-HSCT group and one in MSD-HSCT group died within 21 days, and other patients were engrafted successfully. The median time of neutrophil engraftment was 13 days (10-18 d), 12 days (9-16 d) and 12 days (9-14 d) in Haplo-HSCT + UCB group, MUD-HSCT group and MSD-HSCT group, respectively; the median time of platelets engraftment was 11 days (9-18 d), 12 days (10-23 d) and 12 days (9-14 d), respectively. There were 10, 3, 3 cases of grade Ⅱ-Ⅳacute GVHD at day 100 in the three groups, respectively, and there were 6, 4, 3 cases of chronic GVHD in the three groups, respectively. The 2-year cumulative incidence of relapse was 40.6%, 66.2% and 26.7%, respectively. The predicted 2-year overall survival rate was 37.9%, 42.9% and 55.4%, respectively. All these data had no significant difference (all P＞ 0.05). Conclusion The efficacy of Haplo-HSCT combined with UCB infusion is similar to that of MUD-HSCT or MSD-HSCT in treatment of high-risk lymphoblastic malignancies, which should be recommended to the patients with high-risk lymphoblastic malignancies and without matched donors.