As a transmembrane protein, CD47 is widely distributed in a variety of cells. It can bind to signal regulatory protein alpha (SIRPα) on macrophages and release inhibitory signals, thus avoiding phagocytosis of macrophages. In lymphoma cells, the expression of up-regulation of CD47 expression in lymphoma cells is one of the important mechanisms for inducing immune escape, and it is also a potential therapeutic target. This article reviews the research progress of CD47-induced immune escape, monoclonal antibodies targeting CD47 and cellular immunotherapy in the treatment of lymphoma.

The microenvironment of lymphoma is an important factor affecting the development of lymphoma, which is involved in regulating the recognition and immune response of lymphoma cells by the immune system. In the era of immunotherapy of lymphoma, the state of microenvironment also affects the effect of monoclonal antibodies, small molecular compounds and other immune targeting drugs on lymphoma cells. Among them, microenvironment-related immune escape is one of the key factors leading to the failure of lymphoma treatment. This article reviews some microenvironment factors such as stromal immune cell subsets, vascular proliferation, hypoxia, immune checkpoint and the recent research progress of immune escape.

Objective To explore prevention of cyclosporine A (CsA) combined with Cobalt protoporphyrin (CoPP) against murine graft versus-host disease (GVHD) after allogeneic hematopoietic stem cell transplantation (allo-HSCT).Methods C57BL/6 (H-2Kb) mice were used as donors and BALB/c (H-2Kd) mice as recipients,which were randomly divided into 4 groups.The mice in total body irradiation group (TBI group) were lethally irradiated and injected intravenously with PBS; The mice in Allo-HSCT group (BS group) were lethally irradiated and injected intravenously with bone marrow cells and spleen cells; The mice in CsA intervention group (CsA group) were injected with CsA intraperitoneally after allo-HSCT; The mice in CsA combine with CoPP intervention group (combination group) received both CsA and CoPP intraperitoneally after alloHSCT.Recipients were monitored for condition,survival rate and weight.The liver,small intestine and skin in the recipients were gained and pathological changes of GVHD were assessed.The kidney was stained with Masson staining dye to observe the tissue fibrosis.The expression levels of renal HO-1 mRNA in the recipients were detected.Results In contrast to BS and CsA groups,GVHD degree in combination group was mild,with less reduction and quick recovery of weight.On the day 30 after HSCT,survival rate in BS group was 36.8％,and that in combination group and CsA group was 69.6％ and 53.5％ respectively (P＜0.05).In comparison with BS and CsA groups,pathological changes in combination group were mild,cellular edema and degeneration degree of the liver,small intestine and skin were slight,and few necrosis and infiltrated inflammatory cells were observed.Tubulointerstitial fibrosis hardly occurred in combination group,but it occurred in CsA group abundantly.As compared with BS group,the expression levels of HO-1 mRNA was increased in combination group,while decreased in CsA group (P＜0.05).Conclusion CsA combined with CoPP enhanced the protective effect of CsA against GVHD,moreover,CoPP could alleviate the side effects of CsA,which might be related with up-regulation of the expression levels of HO-1.

Objective To assess the functional role of TH 17 cells in allogeneic hematopoietic stem cell transplantation (allyHSCT).Methods Bone marrow monocytes and splenic T cells were enriched from C57/BL6 donors.Recipient Balb/c mice were irradiated with 7.5 Gy total body irradiation (TBI) and injected with 5 105 splenic T cells and 5 106 bone marrow monocytes.Survival was monitored daily,clinical graft-versus-host disease (GVHD) was assayed three times a week,and detailed histopathologic analyses of lung were performed at day six after Allo-HSCT.Flow cytometry analysis was performed using CD3-FITC,CD4-PE,CD45-PerCP-CyS.5 monoclonal antibodies.Cells were stained for intracellular cytokines using mouse TH 1/TH2/TH 17 cytokine kit.Results All the experimental animals showed GVHD manifestations on the day 6 after transplantation.Animals from BMT and HF groups were scarified and histological analysis of lung was performed.Absence of TH 17 cells induced severe pathologic pulmonary lesions.The histopathology of the lung tissue was characterized by disorganization,epithelia cell damage,interstitial fibroplasias,and monocytes infiltration.The proportion of TH1 and TH 17 in BMT group was (5.53 ± 0.11 ) ％ and ( 1.04 ± 0.34)％ respectively,both significantly different from that in HF group.The levels of IL-17A and IFN-γin BMT group were (2.81 ±0.19) and (42.97 ± 0.23) pg/mL respectively.IL-17A could not be detected in HF group,yet the level of IFN-y was only (9.89 ± 0.51 ) pg/mL.IL-10 in both HF and BMT groups was not detectable.Conclusion Lung is on target of aGVHD.IL-17A may play a key role in the lung injury after transplantation.

Objective To investigate the CD47 expression in de novo acute myelogenous leukemia (AML) patients with normal karyotype and its clinical significance. Methods One hundred thirty-seven cases of de novo AML with normal karyotype and 3 healthy volunteers were selected. Relative CD47 expressions in normal bone marrow hematopoietic stem cells (HSC) and multipotent progenitor (MPP) from healthy volunteers, as well as bone marrow mononuclear cells (MNC) and leukemia stem cells (LSC, Lin-CD34+CD38-CD90-) from AML patients were determined by flow cytometry. CD47 expression on the Lin-CD34+CD38-LSC-enriched fraction of specimen was determined by flow cytometry. The FMS-like tyrosine kinase 3 internal tandem duplication (FLT3-ITD) was detected by using the Genome Analyzer platform. CD34+CD38-CD47hi and CD34+CD38-CD47lo expressing cells were identified and purified using FACS. Two groups of cells were inoculated with MethoCult H4445 medium on agarose-containing methylcellulose plates. After 12 days, MPP colony forming units (CFU) were counted, and 1×105 CD34+ CD38- CD47lo and CD34+ CD38-CD47hi cells were transplanted into NSG (NOD-SCID IL-2R γ null) mice irradiated by 280 cGy, and mice were sacrificed after 8 weeks. The ratio of human CD45+cells was detected by flow cytometry. Results The expression of CD47 in AML patients was higher than that in the healthy control. CD47 was expressed in all FAB (French-American-British) subtypes of AML. No significant difference in CD47 expression among different FAB subtypes was found (F=0.545, P>0.05). Among the 37 patients with CD34+CD38-CD47hi, 17 (46 %) were FLT3-ITD negative, and 20 (54 %) were FLT3-ITD positive. Among the 100 patients with CD34+CD38-CD47hi, 63 (63%) cases were FLT3-ITD negative, 37 (37%) cases were FLT3-ITD positive. The rate of FLT3-ITD positive in patients with CD34+ CD38- CD47lo had no statistical difference compared with patients with CD34+CD38-CD47hi (χ2= 3.79, P> 3.79). The CD34+CD38-CD47lo or CD34+CD38-CD47hi which was selected by FACS, was inoculated with the methylcellulose plate containing agarose for 12 days, and CD34+CD38-CD47lo cells could form CFU. The NSG mouse transplantation experiment showed that CD34+CD38-CD47lo cells could be reconstructed hematopoiesis, and CD34+CD38-CD47hi implantation failed. Conclusion CD34+CD38-CD47hi could enrich LSC, which may be a potential marker to detect minimal residual disease.

Objective To explore the value of microvascular angiography in evaluating the neovascularization of internal carotid artery plaque and the prediction of cerebral infarction.Methods A total of 100 patients with suspected cerebral infarction received from March 19th,2016 to December 1 st,2017 in our hospital were enrolled in this study.All patients underwent microvascular imaging and then analyzed the importance of this technique in evaluating cerebral infarction,plaque neovascularization.Results 100 lesions were recorded in 100 patients,among which there were 22 cases with no superb microvascular ultrasound imaging (SMI) and blood flow signal (ie,score 0) in the plaque.SMI imaging could better show neovascularization (ie score 1 or 2 points).In 78 cases,they all grew from the arterial wall to the plaque.There were 61 cases of cerebral infarction in the blood flow group (78.2％),8 cases (36.3％) of cerebral infarction in the group without blood flow,with statistical significance(P =0.023).SMI imaging was highly sensitive to the prediction of cerebral infarction incidence,reaching 88.4％.Conclusions Ultra-microvascular imaging can improve the sensitivity of cerebral infarction prediction and promptly detect neovascularization of internal carotid artery plaque,thus providing a basis for clinical treatment.

Objective: To construct humanized anti-CD19 chimeric antigen receptor T cells and investigate its ability to kill leukemia cells in vitro and in vivo. Methods: Humanized anti-human CD19 antibody with a high affinity was obtained based on mouse anti-human CD19 antibody (FMC63). Humanized CD19 CAR-T cells (hCART19) were constructed through transfection of lentivirus carrying a CAR sequence of humanized anti-CD19 scFv into human peripheral CD3⁺ T cells. The ability of hCART19 to kill leukemia cells and secrete cytokines was detected by LDH release assay and ELISA. The in vivo tumor-killing effect of hCART19 was evaluated in a leukemia mouse model. Results: Several different humanized CD19 single-chain antibodies which were constructed by IMGT database were expressed in the eukaryotic expression vector and purified followed by acquiring humanized CD19 antibody (Clone H3L2) with similar binding ability to FMC63. Humanized CD19 CAR lentivirus vector was constructed and transfected into T cells to obtain hCART19 cells. The LDH release experiment confirmed that the killing rate of target cells was increased gradually along with the increased E/T ratio. When the ratio of E/T was 10∶1, the killing rate of target cells by hCART19 reached a maximum. When Raji cells were used as target cells, the hCART19 cells group had a significantly higher kill rate [(87.56±1.99)%] than the untransduced T cells group [(19.31±1.16)%] and the control virus transduced T cells group [(21.35±1.19)%](P<0.001). ELISA analysis showed that the secretion of IL-2 [ (10.56±0.88) pg/ml] and IFN-γ [ (199.02±12.66) pg/ml] in the hCART19 cells group were significantly higher than those in the untransduced T cells group [IL-2: (3.55±0.26) pg/ml; IFN-γ: (37.63±0.85) pg/ml] and the control virus transduced T cells group [IL-2: (2.92±0.32) pg/ml; IFN-γ: (52.07±3.33) pg/ml](P<0.001). The above experiments also showed similar results when CHO-K1-CD19 cells were used as target cells. Moreover, in a human leukemia xenograft animal model, the results showed that mice in the untransduced T cells group and the control virus transduced T cells group all died within 20 to 30 days, and the hCART19 cell group survived >40 days, which was more than the survival time of the other two groups of mice. The difference was statistically significant (χ²=11.73, P=0.008). Conclusion Humanized CD19 CAR-T cells with anti-leukemic activity have been successfully constructed, which will lay a foundation for clinical studies in the future.

Objective:To investigate the effect of prognostic nutrition index (PNI) and clinical characteristics on the prognosis of patients with diffuse large B-cell lymphoma (DLBCL).Methods:The clinical data of 236 patients with DLBCL treated in the Affiliated Hospital of Xuzhou Medical University from November 2014 to December 2018 were retrospectively analyzed. X-Tile software and restricted cubic spline (RCS) were used to determine the best cut-off values of PNI, age and hemoglobin; Cox proportional hazard regression model was used for univariate and multivariate analyses; Kaplan-Meier method was used to analyze the overall survival (OS) of patients, and log-rank test was also performed.Results:One-hundred and fifteen of the 236 patients (48.7%) died, with a median OS time of 32 months. The 3-year OS rate was 46%, and the 5-year OS rate was 36%. The best cut-off value of PNI was 49. There was a significant non-linear relationship between PNI and the risk of poor prognosis of DLBCL ( χ2=34.64, P < 0.01); the analysis of the dose-response relationship showed that with the change of PNI, the correlation strength of the risk of poor prognosis declined non-linearly. The best cut-off value of age was 63 years old, and the correlation strength between age and the risk of poor prognosis of DLBCL showed a non-linear upward trend ( χ2=14.86, P=0.022). The best cut-off values of hemoglobin calculated by X-Tile software were 93 g/L and 129 g/L. Multivariate analysis showed that PNI, central nervous system involvement, liver involvement, age, hemoglobin, international prognostic index (IPI) score, and bulky disease were independent influencing factors of OS in DLBCL patients (all P < 0.05). In patients with germinal center B-cell-like (GCB) subtype, bcl-2-positive and bcl-6-positive, there were statistical differences in the 3-year OS rate of patients with PNI < 49 and PNI ≥ 49 (all P < 0.05). Conclusion:PNI has a certain value in the prognosis assessment of DLBCL patients, and PNI ≥ 49 indicates that the patient has a good prognosis.

Objective To evaluate the effect of ditching for drain on the control of the breed of Oncomelania hupensis snails in beaches of Dongting Lake. Methods From November,2009 to November,2012,an O. hupensis snail infested beach of the Yueyang jail and an O. hupensis snail infested beach of Junshan District were selected as research fields in the eastern Dongting Lake area,and the former,as the intervention field,was performed with the ditching for drain by excavators and the latter,as the control field,was not. Results Before the project implemented,the average soil moisture contents on the beaches in dry seasons of the two fields were both about 35.56%. After the project implemented,in the intervention field,the average soil mois?ture content was 26.53%which was significantly lower than that(35.56%)in the control field(F=6.53,P<0.05). The under?ground water levels in different heights in the intervention field were lower than those in the control field (χ2 = 33.33,P <0.05). Before the project implemented,the natural death rates of the snails were 0.98%and 0.89%in the two research fields re?spectively(P>0.05),and after the project implemented(in 2012),no adult and young snails were found in the interventional field,but in the control field,the average densities of living snails and young snails were 29.37 snails/0.1 m2 and 213±108.45 snails/0.1 m2 respectively. Conclusion The intervention of ditching for drain can decrease the soil moisture contents quickly and change the ecological condition,therefore,can control the breed of O. hupensis snails in the beaches of Dongting Lake.

Objective:To explore the influence of prognostic nutritional index (PNI) and controlling nutritional status (CONUT) on the prognosis of patients with multiple myeloma.Methods:Data of 157 patients with multiple myeloma (MM) at the affiliated hospital of Xuzhou medical university from January 2014 to December 2018 were retrospectively evaluated. The operating characteristic (ROC) curve analysis was adopted as the optimal cut-off point. PNI and CONUT were grouped based on the cut-off points of 44.45 and 3.5, respectively, and the differences between age, gender, serum calcium, β 2-microglobulin, serum creatinine, lactate dehydrogenase, and hemoglobin were analyzed. The prognostic factors were analyzed via univariate and Cox multivariate regression analyses. Results:The level of PNI and CONUT is the influencing factor of OS time. The univariate analysis revealed that age, LDH, plasma cell ratio, β 2-microglobulin, ISS stage, PNI, and CONUT were the risk factors for the prognosis of patients with MM. The multivariate analysis revealed that age ( HR=1.636, 95% CI 1.014-2.640) , plasma cell ratio ( HR=1.953, 95% CI 1.232-3.096) , and PNI ( HR=0.513, 95% CI 0.287-0.917) were the independent prognostic risk factors of patients with MM. Conclusion:Low PNI in patients with MM indicates a poor prognosis, which is an independent prognosis risk factor.