Irisin inhibits neutrophil extracellular traps formation and protects against acute pancreatitis in mice.

INTRODUCTION: Irisin is a newly discovered myokine which links exercise to inflammation and inflammation-related diseases through macrophage regulation. However, the effect of irisin on the activity of inflammation related immune cells (such as neutrophils) has not been clearly described. OBJECTIVES: The objective of our study was to explore the effect of irisin on the neutrophil extracellular traps (NETs) formation. METHODS: Phorbol-12-myristate-13-acetate (PMA) was used to construct a classic neutrophil inflammation model that was used to observe the formation of NETs in vitro. We studied the effect of irisin on NETs formation and its regulation mechanism. Subsequently, acute pancreatitis (AP) was used to verify the protective effect of irisin in vivo, which was an acute aseptic inflammatory response disease model closely related to NETs. RESULTS: Our study found that addition of irisin significantly reduced the formation of NETs via regulation of the P38/MAPK pathway through integrin αVß5, which might be the one of key pathways in NETs formation, and which could theoretically offset the immunoregulatory effect of irisin. Systemic treatment with irisin reduced the severity of tissue damage common in the disease and inhibited the formation of NETs in pancreatic necrotic tissue of two classical AP mouse models. CONCLUSION: The findings confirmed for the first time that irisin could inhibit NETs formation and protect mice from pancreatic injury, which further elucidated the protective effect of exercise on acute inflammatory injury.

Elevated serum ferritin levels are associated with severity and prognosis of severe acute pancreatitis: a preliminary cohort study.

BACKGROUND: Serum ferritin (SF), as an acute-phase response protein, is used to reflect the degree of oxidative stress and systemic inflammatory responses. This study was designed to assess the effect of elevated SF levels on the severity of acute pancreatitis (AP). METHODS: From January 2013 to December 2020, 200 consecutive patients with AP were retrospectively reviewed to analyze the relationships among the etiologies of pancreatitis, the severity of the disease and SF levels. The receiver operating characteristic (ROC) curve and logistic regression analysis were used to assess whether elevated SF levels could predict the onset of organ failure in AP. RESULTS: 92 (46%) had high SF levels (> 275 ng/ml). SF levels were not associated with the etiology of AP disease. Among patients with high SF levels, there was a significant increase in the proportion of patients with severe AP (23.1% vs. 76.9%) and a higher proportion of systemic inflammatory response scores (25.9% vs. 44.6%) in comparison to patients with normal SF levels. The area under the ROC curve for SF in predicting persistent organ failure was 0.812 [95% confidence interval 0.721-0.904]. CONCLUSIONS: F concentrations were positively correlated with the severity of AP, and quantitative assessment of SF can predict disease severity and organ failure in patients with AP.

Feasibility and effectiveness of prone position ventilation technique for postoperative acute lung injury in infants with congenital heart disease: study protocol for a prospective randomized study.

BACKGROUND: Prone position ventilation is a widely used lung protection ventilation strategy. The strategy is more convenient to implement in children compared to adults. Due to the precise mechanism of improving oxygenation function, development of pediatric prone ventilation technology has been largely focused on children with acute respiratory distress syndrome. There is a paucity of high-quality studies investigating the effects of prone position ventilation after pediatric cardiac surgery. The purpose of this study is to evaluate the feasibility and effectiveness of prone position ventilation in infants who develop postoperative acute lung injury after surgery for congenital heart disease. METHODS: A single-center, randomized controlled trial of pediatric patients with acute lung injury after surgery for congenital heart disease who will receive prone position ventilation or usual care (control group). A total of 68 children will be enrolled according to the inclusion criteria. The main outcome measures will be lung compliance and oxygenation index. The secondary outcomes will be duration of mechanical ventilation, length of stay in cardiac intensive care unit, reintubation rate, and complication rate. DISCUSSION: This study will investigate the feasibility and effectiveness of prone position ventilation techniques in children who develop postoperative acute lung injury after surgery for congenital heart disease. The results may help inform strategies to improve airway management after surgery for congenital heart disease. TRIAL REGISTRATION: ClinicalTrials.gov NCT04607993 . Initially registered on 29 October 2020.

Impact of Fatty Pancreas on Postoperative Pancreatic Fistulae: A Meta-Analysis.

BACKGROUND: Soft pancreas is widely recognized as an important risk factor for the development of postoperative pancreatic fistula (POPF). Although fatty pancreas (FP) has not been formally defined as a cause of pancreatic fistula, existing research has shown that it can increase the incidence of POPF by increasing pancreatic tenderness; therefore, it may be a potential risk factor. This study aimed to discern whether FP was associated with POPF. METHOD: Two reviewers independently performed literature searches from five electronic databases. According to the established inclusion criteria, we extracted necessary data from the studies that met the criteria for further analysis. We pooled the odds ratios (ORs) from individual studies using a random-effects model to investigate the associations between POPF and the prognosis of FP. RESULT: A total of 11 studies involving 2484 individuals were included. The pooled prevalence of POPF was 18% (95% CI: 12-24%). Body mass index (BMI) was associated with a significantly increased risk of POPF (OR=3.55; 95% CI: 1.83, 6.86; P=0.0002; I²=0). FP was obviously associated with the occurrence of POPF (OR=3.75; 95% CI: 1.64, 8.58; P=0.002; I²=78). CONCLUSION: FP is closely associated with the development of POPF, and the early identification of these high-risk patients can help to reduce the incidence of POPF. SYSTEMATIC REVIEW REGISTRATION: The Registration URL link is (https://www.crd.york.ac.uk/PROSPERO/). The ID is "CRD42021265141".

The mediating effect of self-efficacy on the relationship between family functioning and quality of life among elders with chronic diseases.

AIM: To explore whether self-efficacy has any positive or negative mediating effects between family functioning and quality of life among elders with chronic diseases. DESIGN: A cross-sectional study. METHODS: Questionnaires were collected from 516 community-dwelling elderly individuals with chronic diseases using a convenience sampling method. The questionnaires included the Self-efficacy for Managing Chronic Disease Six-Item Scale, the Family Adaptation Partnership Growth Affection Resolve Index and the MOS 36-Item Short Form Health Survey. RESULTS: Family functioning and self-efficacy impacted the quality of life of community-dwelling elderly individuals with chronic diseases. Family functioning was mediated by self-efficacy and had an indirect impact on quality of life. The mediating effect accounted for 62.50% of the total effect.

Postgraduate nursing internationality education: A study in establishing an English nursing course.

Aim: The study was to explore the significant influence of an English nursing course in nursing postgraduate internationalization education. Design: A cross-sectional study. Method: The research object included three grades (a total of 18) of nursing postgraduate students in the Nursing School of Yangzhou University. A standardized four-section questionnaire designed by the authors was applied to the survey. Results: 88.89% were satisfied with the course design and application of the English nursing course, and the scores for all 6 items were above the average, but only 44.4% of the postgraduate students understood the course completely. In teaching design and content aspect, 88.9%-94.4% postgraduate nurses felt that the course learning requirements were clear and could improve their knowledge and capacity for scientific research. All of the postgraduate nurses were identified with the teacher's moral and academic characters. Conclusions: To establish English nursing courses for nursing postgraduate students is beneficial for postgraduate internationalization education.

Peroxiredoxin 1 as an inflammatory marker in diarrhea-predominant and postinfectious irritable bowel syndrome.

BACKGROUND: Low-grade inflammation occurs in some patients with irritable bowel syndrome (IBS). However, the exact inflammatory markers of IBS and the relationship of these markers with IBS subtypes and symptoms are poorly defined. Peroxiredoxin 1 (PRDX1) plays an important role in inflammatory responses, including intestinal inflammation. We investigated whether PRDX1 is associated with the diagnosis, subtypes, and symptom severity of IBS. METHODS: A total of 177 IBS patients and 174 sex- and age-matched healthy controls (HCs) were recruited. The PRDX1 levels in the sera and colonic mucosa of the participants were detected by enzyme-linked immunosorbent assays and immunohistochemistry. The severity of IBS symptoms was assessed using the IBS Severity Scoring System (SSS) questionnaire. RESULTS: The PRDX1 levels in the sera (F = 71.81, P < .001) and colonic mucosa (F = 5.359, P < .001) of postinfectious (PI-IBS) and diarrhea-predominant IBS (IBS-D) groups were significantly higher than those of the other three IBS subtypes and HC group. The PRDX1 level in the serum and colonic mucosa of IBS-D (serum, P < .01, mucosa, P < .001) and PI-IBS (serum, P < .05, mucosa, P < .001) groups with the most severe symptoms was significantly higher than that in the groups with mild and moderate symptoms. Correlation analysis revealed that in patients with IBS-D (P < .001) and PI-IBS (P < .05), the levels of PRDX1 and TNF-α in sera had a significant positive correlation with IBS-SSS. CONCLUSION: Elevated PRDX1 in the serum and colon mucosa may be closely related to the progression of IBS (especially IBS-D and PI-IBS) and the expression of gastrointestinal symptoms.

[Advance in studies on antitumor and immunomodulatory effects of wogonin].

Wogonin is a kind of natural flavonoid compound. According to findings in the latest studies, wogonin shows a wide range of antitumor effects, with the characteristics of multi-pathway, multi-link and multi-target, such as promoting tumor cell apoptosis through ROS or Ca(2+)-mediated signal paths, enhancing tumor cytotoxicity by TNF-α and TRAIL, blocking tumor cell cycle, inhibiting tumor angiogenesis and resisting cancer synergistically with chemotherapeutic drugs. Moreover, Wogonin could enhance body immune function by enhancing immune cell infiltration, regulating the immune cell phenotype and promoting relevant cytokine secretion. In this paper, the authors summarized the advance in studies on wogonin's antitumor and immunomodulatory effects.

IL-15, in synergy with RAE-1ɛ, stimulates TCR-independent proliferation and activation of CD8(+) T cells.

CD8(+) T cells play critical roles in immunosurveillance by killing malignant or virally infected cells. Interleukin 15 (IL-15) is a critical cytokine for promoting proliferation and the effector capacity of CD8(+) T cells, and has been used to support the growth of CD8(+) T cells in cellular therapies of neoplastic diseases. Recent studies have shown that IL-15, in synergy with other cytokines, such as IL-6, enhances the T-cell receptor (TCR)-independent proliferation and function of CD8(+) T cells. The aim of the present study was to investigate the role of BaF3-mb15-RAE cells in stimulating mouse CD8(+) T cells. BaF3 cells were cultured and B16F10 cells were grown in DMEM. MTT assay was used to detect the proliferation of CD8(+) T cells. Cells were analyzed using flow cytometry. The results showed that IL-15 synergistically acts with another T-cell stimulatory molecule, RAE1ɛ, to potently promote the proliferation of CD8(+) T cells, induce CD8(+) T-cell activation and enhance granzyme B and interferon-γ (IFN-γ) production in the absence of signaling via the TCR. Moreover, IL-15 in combination with RAE1ɛ resulted in a cooperative effect on CD8(+) T-cell-mediated cytotoxicity against B16F10 tumor cells. Thus, results of the present study showed that IL-15, in synergy with RAE1ɛ, enhances the TCR-independent effector function of CD8(+) T cells in vitro, which may be useful in the cellular immunotherapy of cancer.

[Effects of natural killer cells stimulated with immobilized MHC class I chain-related antigen A on activities of dendritic cells].

AIM: To observe the effects of NK cells stimulated with immobilized MHC class I chain-related antigen A (iMICA) on activities of dendritic cells (DCs). METHODS: Firstly fresh allogeneic NK cells, or iMICA-stimulated allogeneic NK cells, and autologous NK cells stimulated with IL-2 or IL-2 and iMICA were co-cultured with immature DCs (iDCs) at the ratio of 5:1 for 24 hours. Frequencies of HLA-DR positive or CD86 positive DCs were detected by flow cytometry. Next autologous NK cells stimulated with iMICA were co-cultured with iDCs at the ratio of 1:5 for 24 hours. Variation of HLA-DR or CD86 expression on dendritic cells was measured. Lastly IFN-γ neutralizing antibody was added into the NK-DCs co-culture system to observe HLA-DR or CD86 expression on DCs. RESULTS: When NK:iDCs ratio was 5:1, both fresh allogeneic and activated autologous NK cells killed iDCs efficiently. iMICA did not synergize this cytotoxicity. However, when NK:iDCs ratio was 1:5, NK cells activated by iMICA promoted HLA-DR and CD86 expression on DCs. IFN-γ antibody down-regulated HLA-DR and CD86 expression on DCs which were co-cultured with iMICA-activated NK cells. CONCLUSION: iMICA is redundant as fresh allogeneic or activated autologous NK cells killed iDCs. However, if numbers of NK cell are less than those of DCs, iMICA can stimulate NK cells to produce IFN-γ for DCs maturation.

Construction of a plasmid for co-expression of mouse membrane-bound form of IL-15 and RAE-1ε and its biological activity.

Interleukin 15 (IL-15) is a pivotal cytokine for the proliferation and activation of a specific group of immune cells such as natural killer (NK), IFN-producing killer dendritic cells (IKDC) and CD8 T cells. RAE-1ε, the ligand for the activating NKG2D receptor, which also play an important role in the proliferation and activation of NK cells and IKDCs. In this study, a membrane-bound form of IL-15 (termed mb15) encoding sequence and RAE-1ε gene were obtained by SOE-PCR or PCR amplification. The amplified mb15 and RAE-1ε gene were then digested and inserted into the multiple cloning site1 (MCS1) and MCS2 of pVITRO2-mcs vector, respectively. A recombinant eukaryotic expression vector for co-expression of mb15 and RAE-1ε was successfully constructed. After it was transfected to BaF3 cells, the expression of IL-15 and RAE-1ε in recombinant BaF3/mb15/RAE-1ε cells were verified by RT-PCR, western blot and FCM analysis. Furthermore, BaF3/mb15/RAE-1ε cells had the ability of promoting NK cells proliferation and IFN-γ secretion. In conclusion, BaF3/mb15/RAE-1ε cells were successfully constructed, which is very useful for further studies, especially for the expansion and activation of certain subsets of immune cells such as NK cells and IKDCs.

[The effect of up-regulation of MHC class I chain-related antigen A expression by dendritic cells on activity of NK cells].

AIM: To observe whether MHC class I chain-related antigen A (MICA) was expressed on monocytes, immature dendritic cells (iDCs), and mature dendritic cells (mDCs), and to study effect of up-regulation of MICA expression by DCs on biologic activity of NK cells. METHODS: MICA expression on monocytes, iDCs, or mDCs stimulated with LPS, TNF-α, CD40L, IL-15 or IFN-α was detected by flow cytometry. Next CD69 expression, degranuation, and IFN-γ production of NK cells stimulated with MICA-positive mDCs were analyzed. Lastly recombinant NKG2D/Fc fusion protein and anti-IL-12 monoclonal antibody was respectively added into culture systems to analyze whether these reagents affected the interaction between DCs and NK cells. RESULTS: MICA was not expressed on monocytes, and expressed on iDCs at low level. LPS, TNF-α, CD40L had no influences on MICA expression on mDCs, but IFN-α, IL-15 up-regulated MICA expression on mDCs. MICA-positive mDCs promoted CD69 expression, IFN-γ production, and killing K562 cells by NK cells. NKG2D/Fc inhibited both cytotcoxicity and IFN-γ secretion, whereas IL-12 antibody only inhibited IFN-γ secretion of NK cells. CONCLUSION: MICA expression on DCs is regulated by relevant factors in microenvironment. DCs with high level of MICA expression can up-regulate biologic activity of NK cells.

[Association of MICA gene polymorphism and serum soluble MICA level with colorectal cancer].

OBJECTIVE: To investigate whether the major histocompatibility complex class I chain-related gene A gene (MICA) polymorphism and serum soluble MICA level were associated with the occurrence and development of colorectal cancer. METHODS: DNA samples from 117 colorectal cancer patients and 113 healthy individuals from Yangzhou in Jiangsu province were genotyped by using the polymerase chain reaction (PCR) and sequence-specific primer (SSP) method and PCR based sequencing. In addition, polymorphism at position 129 was also analyzed by PCR-SSP. Serum levels of soluble MICA were measured by a sandwich ELISA method. RESULTS: Neither the extracellular nor the transmembrane region polymorphisms of MICA gene were associated with the occurrence and the different stages of colorectal cancer. In contrast, the frequency of the methionine residue at position 129 was significantly decreased in the patient group. Soluble MICA levels in sera were increased in the late stages of colorectal cancer. CONCLUSION: Although there was no genetic susceptibility attributed to MICA gene polymorphism with regard to development of colorectal cancer, serum levels of soluble MICA may be a diagnostic marker of advanced stages.

[Identification of T cell epitopes of p210(bcr-abl); antigen and detection of epitope-specific CTLs.].

AIM: To study the immunogenicity of p210(bcr-abl);, the epitopes of HLA-A2 restricted T cells and the distribution of epitope-specific CTLs in chronic myeloid leukemia (CML) patients and in normal controls. METHODS: Two epitopes, BCR-ABL(642); and BCR-ABL(926m);, were selected using bioinfomatics software and further confirmed by T2 cell binding assay. The soluble HLA-A2 tetramers bound with each epitope were generated to detect the CD8(+); T cell frequencies in peripheral blood mononuclear cells. RESULTS: The epitope-specific CD8(+); T cell frequencies of both BCR-ABL(642); and BCR-ABL(926m); were significantly higher in CML patients, compared with those in healthy individuals(P<0.01), but no significant difference was observed in influenza epitope-specific CTLs between the patients and healthy individuals (P>0.05). The frequency of BCR-ABL(642); peptide-specific CTLs at chronic phase was significantly higher than that at blast phase in CML patients (P<0.05). CONCLUSION: The two candidate epitopes selected from p210(bcr-abl); are characterized by their immunogenicity and based on them, vaccines or adoptive CTL therapies can be developed.

[Effects of recombinant soluble MICA protein on the biologic activities of NK cells].

AIM: To study the effects of recombinant soluble MHC class I chain-related protein A (sMICA) on the cytotoxicity, secretion of IFN-gamma, proliferation and apoptosis of peripheral NK cells. METHODS: After NK cells were co-cultured with recombinant soluble MICA proteins overnight, the cytotoxicity of NK cell on target cells was detected by flow cytometry. The supernant was collected to determine the concentration of IFN-gamma by ELISA. The proliferation of NK cells to sMICA was detected by MTS/PMS. NK cells were labeled with annexin V and PI to analyze their apoptosis. RESULTS: Soluble MICA inhibited the cytotoxicity of NK cells and down-regulated the secretion of IFN-gamma, but it showed no effects on the proliferation and apoptosis of freshly isolated peripheral NK cells. CONCLUSION: The soluble MICA shedding from tumor cells could be a pathway of cancer immune evasion by down-regulating the biologic activities of NK cells.

[Correlation of Helicobacter pylori infection with the expression of COX-2 and EGFR and VEGF in human gastric carcinoma].

OBJECTIVE: To investigate the correlation of Helicobactor pylori (Hp) infection with the expression of COX-2, EGFR and VEGF in human gastric carcinoma. METHODS: The expression of COX-2, EGFR and VEGF was detected by immunohistochemistry in samples of 61 gastric cancers and 20 cancer-adjacent tissues. Western blotting was performed in samples of 10 gastric cancers and corresponding cancer-adjacent tissues. Hp infection was detected in 47 patients by fast urea enzyme test and (13)C breath test. RESULTS: The results of immunohistochemistry showed that the positive expressions of COX-2, EGFR and VEGF in gastric carcinoma were 59.02%, 36.07% and 60.66%, respectively, significantly higher than that in the normal mucosa (25.00%, 0 and 30.00%, respectively). There was a positive correlation between the expression of COX-2, EGFR and VEGF and gastric carcinoma. The expression of COX-2 and EGFR was 75.76% and 45.45% in the gastric carcinomas with Hp infection, significantly higher than that in those without (28.57% and 14.29%). The protein expression of COX-2, EGFR and VEGF detected by Western blot in gastric carcinomas was also significantly higher than that in normal mucosa. CONCLUSION: COX-2, EGFR and VEGF are overexpressed in gastric carcinoma, and there is a positive correlation among them. Hp infection may upregulate the expression of COX-2 and EGFR in gastric cancer tissues.

[Immune protective mechanisms of gene vaccines with co-expressing bcr-abl fusion gene fragment and mouse IL-7 gene].

OBJECTIVE: To investigate the influence of mIL-7 on the immune response induced by vaccine of bcr-abl fusion gene fragment in mouse. METHODS: BALB/c mice were immunized by i. m. injection of pVbcr-abl/mIL-7 and pVbcr-abl, respectively. The specific antibody to p210bcr-abl protein was assayed by ELISA. The CTL activity of spleen cells from the immunized mice was assessed with LDH release test. RESULTS: The pVbcr-abl/mIL-7 and pVbcr-abl-immunized BALB/c mice elicited higher specific antibodies to p210bcr-abl protein. The specific antibody level of former group was higher than that in latter group, but the difference was statistically not significant. The spleen cells from the immunized mice showed more effective CTL activity than that from control group. The cytotoxic activity of spleen CTLs induced by pVbcr-abl/mIL-7 immunized mice exceeded that of pVbcr-ab-immunized mice. CONCLUSION: The mIL-7 may influence the growth and differentiation of T cells, promote some T cells migrating into tumor tissue and up-regulate the specific cellular immune response. The results of this study provided an useful experimental basis for preclinical research on gene vaccine for chronic myeloid leukemia.

[Influences of bcr-abl gene vaccine on inoculated SP2/0/bcr-abl tumor cells in mice].

To study the influence of vaccine of bcr-abl fusion gene fragment on inoculated SP2/0/bcr-abl tumor cells in mice, BALB/c mice were immunized with pVbcr-abl, pVbcr-abl/mIL7 plasmids, respectively, then SP2/0/bcr-abl cells expressing the fragment of bcr-abl fusion gene were inoculated subcutaneously into the groin of BALB/c mice in order to observe the effect of vaccine on growth of inoculated SP2/0/bcr-abl tumor cells. The results showed that there were distinct differences on the time of tumor growth, the time of tumor ulceration, tumor volume and survival time of mice bearing tumor between two immunized groups and two control groups (blank and vacant plasmid groups). The mice immunized with pVbcr-abl/mIL7 lived longer as compared to mice immunized with pVbcr-abl. The tissue of inoculated tumor was more compact, tumor organ was larger, tumor form was irregular in 2 control groups, while the tissue of inoculated tumor was looser, tumor volume was smaller, and with mass inflammatory infiltration in two immunized groups. Moreover, the metastatic tumor cells were found in the livers of control groups, but not observed in two immunized groups. It is concluded that the protection occurred in immunized mice which inhibited the growth of SP2/0/bcr-abl tumor cell in vivo.

[Recombinant eukaryotic expression plasmid of bcr-abl gene fragment induces specific immune response in mice].

OBJECTIVE: To study the specific immune response induced by a recombinant eukaryotic expression plasmid encoding bcr-abl fusion gene fragment so as to explore new immunotherapy in mouse. METHODS: A recombinant eukaryotic vector pVbcr-abl expression cDNA fragment of bcr-abl fusion gene was constructed and used to immunize BALB/c mice. Serum level of bcr-abl specific antibody was detected by enzyme-linked immunosorbent assay (ELISA). Twenty days later the immunized mice were subcutaneously inoculated SP2/0/bcr-abl cells. The survival time, tumor growth time and lymphocytic infiltration were observed. T cells infiltration into tumor tissue was analyzed by immunohistochemistry. Changes of T cell subset in the spleen of mice was analyzed by fluorescent-activated cell sorting (FACS) and the cytotoxicity T lymphocyte (CTL) activity in spleen by lactate dehydrogenase (LDH)-release assay. RESULTS: The eukaryotic expression vector pVbcr-abl was constructed successfully, and highly expressed the cDNA fragment of bcr-abl fusion gene. The BALB/c mice immunized with the vector could generate the specific antibody and CTL, resulting in a specific immunoprotection. There were dramatic differences in the tumor-forming time, tumor ulcer appearing time and tumor-growing speed between the immunized and the control groups. The mice had longer survival time in the immunized group than in the control group. There were a large amount of CD3(+) T cells infiltration in tumor tissue of the immunized mice. The spleen cells from the immunized mice had higher CTL activity with a alteration of T cell subset, the CD4(+)/CD8(+) ratio being 1.54 +/- 0.29, higher than that of control group (1.18 +/- 0.30). CONCLUSION: The recombinant eukaryotic expression plasmid pVbcr-abl can induce in vivo not only the generation of specific antibody, but also high level of specific CTL activity, resulting in killing the SP2/0/bcr-abl tumor cells directly and inhibiting the tumor growth.

[Establishment of mouse SP2/0 cell line stably expressing bcr-abl fusion gene fragment].

To establish SP2/0 cell line H-2(d) stably expressing bcr-abl fusion gene fragment, the bcr-abl fusion gene was subcloned into retroviral vector pLXSN from pGEMbcr-abl. The recombinant retroviral vector pLXSNbcr-abl was transfected into PT67 packaging cells with the help of lipofectamine. The positive clones were selected out and cultured after G418 selection. Then viral supernatant was collected to determine viral titer, the viral titer was 2 x 10(7) CFU/ml. The SP2/0 cells were infected with the collected viral supernatant. The results showed that after G418 selection, the bcr-abl fusion gene was integrated into the chromosome of SP2/0 cells infected stably, with recombinant retrovirus and expressed in SP2/0 cells confirmed by PCR and RT-PCR respectively. In conclusion, the mouse tumor cell lines expressing the bcr-abl fusion protein were successfully established and would be used as a experimental cell model for anti-CML immunotherapy.