Objective:To compare the clinical efficacy between radiofrequency ablation(RFA)and pulmonary metastasectomy in the colorectal cancer(CRC)patients with lung metastases after radical resection. Methods:The clinical data of 80 CRC patients with lung metastases after radical resection were analyzed retrospectively,and were divided into the surgery group(33 cases)and the RFA group(47 cases)according to the local treatment.The overall survival(OS)and progression-free survival(PFS)of the two groups were compared,as well as the prognostic factors of patients were analyzed. Results:The 3-year PFS and OS rates were 42.4％vs 31.9％and 75.8％vs 72.3％in the surgery group and the RFA group,respectively.There was no significant difference in PFS and OS between the two groups(P＞0.05).In multivariate analysis,maximum lung metastasis diameter,preoperative serum carcinoembryonic antigen(CEA)level and history of extrapulmonary metastasis were independent factors influencing OS in the CRC patients with lung metastases after radical resection(P＜0.05).In addition,preoperative serum carcinoembryonic antigen(CEA)level and history of extrapulmonary metastasis were also independent factors influencing PFS in the CRC patients with lung metastases after radical resection(P＜0.05). Conclusion:The short-term efficacy of RFA is comparable to that of pulmonary metastasectomy in the CRC patients with lung metastases after radical resection,and long-term follow-up studies are needed.

Objective:To summarize the clinical, pathological and muscle magnetic resonance imaging (MRI) features of human immunodeficiency virus (HIV)-associated nemaline myopathy (NM; HIV-NM).Methods:The present patient was a 23-year-old man with HIV infection who developed progressive proximal weakness and atrophy for more than 10 months. He was admitted to the Department of Neurology of Beijing Ditan Hospital in early June 2021. Electromyography showed myogenic findings. The serum creatine kinase was 202.4 U/L. CD 4+ count was 585×10 6/L. Serum monoclonal immunoglobulin (M protein) was negative. The patient underwent MRI examination of bilateral thigh muscles, biopsy of left biceps brachii and gene detection. The clinical, pathological and muscle MRI changes of HIV-NM were summarized based on the literature review. Results:MRI examination of bilateral thigh muscles showed edema changes. Muscle biopsy showed nemaline structures in some muscle fibers, accompanied by fiber atrophy and regeneration. No gene mutation related to clinical phenotype was found by second-generation sequencing. After intravenous immunoglobulin combined with prednisone, the patient′s weakness symptoms were significantly improved. A total of 17 cases of HIV-NM (including the present case) were identified in the literature, who were aged (33.7±9.1) years. Fifteen were males and two were females. All patients developed proximal limb weakness. Creatine kinase was normal or slightly elevated. Serum monoclonal protein was positive in 3 cases (3/7). Immunosuppressive therapy was effective.Conclusions:The main clinical characteristics of HIV-NM are progressive proximal limb weakness and muscle atrophy. The muscle pathology shows a large number of nemaline structures in atrophic muscle fibers. Muscle edema can be seen on muscle MRI. This is the first report of HIV-NM in China, which may be a special subtype of immune myopathy.

Objective:To investigate the characteristics and cytotoxicity in vitro of the residual leukemia cells in the culture system that caused the accidental transfer of CD19 chimeric antigen receptor (CAR) into leukemia cells during the preparation of autologous CD19 CAR-T cells of relapsed/refractory B-cell acute lymphoblastic leukemia.Methods:①Peripheral blood mononuclear cells (PBMC) of 30 patients with relapsed/refractory B-cell acute lymphoblastic anemia (R/R B-ALL) who accepted CD19 CAR-T cell therapy and six healthy volunteers were collected. ②The residual leukemia cells were analyzed by flow cytometry in the system after the PBMCs of R/R B-ALL patients were sorted by CD3 magnetic beads. ③ CD3 + T cells from patients and healthy volunteers were transfected with CD19 CAR and CD22 CAR lentivirus to prepare CD19 CAR-T and CD22 CAR-T cells. ④The Nalm-6 cell line was resuscitated and the Nalm-6 cells with CD19 CAR lentivirus were transfected to prepare CD19 CAR-Nalm-6 cells. The patient's primary ALL cells were transfected with CD19 CAR lentivirus at the same time. ⑤The transfection rates were analyzed by flow cytometer, the cell proliferation was analyzed by the CCK-8 method, and the cell-killing activities were detected by the lactate dehydrogenase method. Results:① Among the 30 R/R B-ALL patients who received CD19 CAR-T cell therapy, two patients had 2.04% and 3.32% residual leukemia cells in CD3 + T cells. After 4 days in culture, the residual leukemia cells disappeared and could not be detected by a flow cytometer with prolonged cultivation in vitro. ② The proliferation of CD19 CAR-Nalm-6 cells was higher than that of the Nalm-6 cells. ③ The killing activity of the CD19 CAR-T cells on Nalm-6 cells was higher than that of the CD19 CAR-Nalm6 cells at a target ratio of 1∶1 on 24, 48, 72 h, respectively. The cytotoxicity of CD22 CAR-T cells on CD19 CAR-Nalm-6 cells was significantly higher than that of CD19 CAR-T cells. ④ The cytotoxicity of CD22 CAR-T alone on CD19 CAR-Nalm-6 cells was higher than that of CD19 CAR-T combined with CD22 CAR-T at the same target ratio. Conclusion:The residual leukemia cells in the culture system in the preparation of CD19 CAR-T cells may lead to the introduction of CD19 CAR into leukemia cells and results in the failure of the CD19 CAR-T cell therapy. Detecting the residual leukemia cells in the culture system via flow cytometry before transfection with CD19 CAR lentivirus is needed. Thus, CD22 CAR-T cell therapy could be used as one of the salvage treatments.

Objective: To observe the changes of PD-1 expression, mRNA level and cytotoxic activity of CD19 CAR-T cells during the culture process of CAR-T cells. Methods: The peripheral blood T cells of 6 lymphoma patients with high expression of PD-1 and 6 healthy volunteers were the source of CAR-T cells. The expression of PD-1 was analyzed by flow cytometry. The mRNA level of PD-1 was analyzed by PCR. The cell proliferation was analyzed by CCK-8 assay. The cytotoxicity was analyzed by LDH assay. Results: ①The transfection efficiency of high PD-1 expression T cells and healthy volunteer T cells were as the same (P>0.05) . ②The cell proliferation capacity of CD19 CAR-T cells from high PD-1 expression T cells or healthy volunteer T cells, with or without PD-1 inhibitor were as the same (P>0.05) . ③The cytotoxicity to lymphoma cells of high PD-1 expression T cells and CAR-T cells were lower than that of these two T cells combined with PD-1 inhibitor and the CAR-T cells from healthy volunteer T cells (P<0.001) . There was no difference of the cytotoxicity between the CAR-T cells from high PD-1 expression T cells combined with PD-1 inhibitor and the CAR-T cells from healthy volunteer (P>0.05) . ④There was no difference of the expression of PD-1 in all CAR-T cell groups during the culture process (P>0.05) . There was no difference of mRNA level of PD-1 in all groups during the culture process (P>0.05) . ⑤The PD-1 expression of CAR-T cells increased by the time of culture after contacting with lymphoma cells (P<0.001) . The PD-1 inhibitors could antagonize this effect. There was no difference of mRNA level of PD-1 in all groups after contacting with lymphoma cells (P>0.05) . Conclusion The PD-1 expression of CAR-T cells from high PD-1 expression T cells increased by the time of culture after contacting with lymphoma cells. However, the mRNA level of PD-1 of all groups did not change, even if PD-1 inhibitor was applied.

Objective To investigate the distribution characteristics and risk factors of intracranial and extracranial aterial lesions in Chinese patients with ischemic stroke . Methods In this multi‐center study ,2 310 continuously inpatients with ischemic stroke diagnosed in 20 stroke screening and prevention project base hospitals from June 2015 to M ay 2016 were enrolled . Carotid ultrasonography and transcranial color‐coded sonography or transcranial Doppler were performed in all patients to confirm the presence of cerebral artery stenosis or occlusion . According to the distribution of lesions ,the subjects were divided into 2 groups :the simple intracranial artery stenosis group and the simple extracranial artery stenosis group . T he difference of risk factors between the two groups was compared . Results Of the 2 310 patients with ischemic stroke ,1 516 ( 65 .6% ) had simple intracranial artery stenosis and 794 ( 34 .4% ) had simple extracranial artery stenosis . T he incidence of anterior circulation artery stenosis was higher in the group of intracranial artery stenosis than that in the extracranial artery stenosis group ( 68 .1% vs 48 .7% , P ＜0 .001) . Posterior circulation artery stenosis and combined anterior with posterior circulation artery stenosis were more common in patients with extracranial artery stenosis group than those in intracranial artery stenosis group ( 36 .4% vs 22 .1% ,14 .9% vs 9 .8% ;all P ＜0 .001) . Univariate analysis of risk factors for stroke showed that patients with intracranial arterial stenosis had a higher prevelence of hypertension , diabetes ,obesity ,and family history of stroke ,and their systolic blood pressure ,diastolic blood pressure , body mass index ( BM I) ,fasting blood‐glucose ,glycosylated hemoglobin ,triacylglycerol ,total cholesterol , and low‐density lipoprotein cholesterol were significantly higher than those in the extracranial arterial stenosis group ( all P ＜ 0 .05 ) . T he proportion of elderly ( ≥ 65 years old ) ,male and smokers in the extracranial arterial stenosis group was significantly higher than that in the intracranial arterial stenosis group ( all P ＜0 .05) . Multivariate logistic regression analysis showed that elderly ( ≥65 years old) ,male , and smoking history were independent risk factors for extracranial arterial stenosis ( OR＝ 2 .012 ,1 .637 , 1 .325 ,respectively ;all P ＜0 .05) . While hypertension ,diabetes ,less physical activity ,and high BM I levels were independent risk factors for simple intracranial arterial disease ( OR ＝ 1 .301 ,1 .252 ,1 .248 ,1 .030 , respectively ;all P ＜0 .05) . Conclusions There are significant differences in the distribution characteristics and risk factors of intracranial and extracranial aterial lesions in patients with ischemic stroke in China .

Objective: To investigate the efficacy and safety of CD19 chimeric antigen receptor T (CAR-T) lymphocytes for the treatment of B cell lymphoma. Methods: A total of 22 patients with B-cell lymphoma from February 1, 2017 to July 1, 2018 were reviewed to evaluate the efficacy and adverse reactions of CD19 CAR-T. Results: Of 22 patients with B-cell lymphoma received CD19 CAR-T cells, the median dose of CAR-T cells was 7.2 (2.0-12.0) ×10⁶/kg. Nine of 12 cases of relapse refractory patients were overall response. Complete remission (CR) occurred in 2 of 12 patients, partial remission (PR) in 7 of 12 patients. The overall response in minor residual disease positive (MRD) group was 8 of 10 patients. CD19 CAR-T cells proliferated in vivo and were detectable in the blood of patients. The peak timepoints of CAR-T cells proliferated in the relapsed refractory and MRD positive groups were 12 (5-19) and 4.5 (1-12) days after treatment respectively, and among peripheral blood cells, CAR-T cells accounted for 10.10% (3.55%-24.74%) and 4.02% (2.23%-28.60%) of T lymphocytes respectively. The MRD positive patients achieved sustained remissions during a median follow-up of 8 months (rang 3-18 months) . None of all the patients relapsed during a median follow-up time of 10 months (3-18 months) . However, 7 PR responders of the relapsed refractory patients maintained a good condition for 1.5-6.0 months. One patient bridged to hematopoietic stem cell transplantation, another one sustained remission for 12 months. Cytokine-release syndrome (CRS) occurred in 14 patients with grade 1-2 CRS in MRD positive group and grade 3 CRS in relapsed refractory group. Conclusions CAR-T cell therapy not only played a role in the rescue treatment of relapsed and refractory patients, but also produced a surprising effect in the consolidation and maintenance of B-cell lymphoma. CD19 CAR-T cells might be more effective in the treatment of MRD positive B-cell lymphoma patients than in the refractory or relapsed cases. High response rate was observed with fewer adverse reactions.

Objective: To retrospectively analyze the efficacy and safety of modified cell infusion method in reducing the incidence of febrile non-hemolytic transfusion reaction (FNHTR). Methods: A total of 69 patients were enrolled in the clinical trial of CD₁₉ chimeric antigen receptor T (CAR-T) cell treatment from February 2017 to October 2018. Study group received the modified cell infusion method, that 1×10⁶ CAR-T cells were re-suspended in 2 mg human serum albumin with total volume of 20 ml and injected intravenously. The control group was intravenously administrated with CAR-T cell in 100 ml normal saline. The incidence of FNHTR, cytokine releasing syndrome (CRS) grade, cytokine level and efficacy were compared. Results: (1)The incidence of FNHTR in the study group was 21.1%, significantly lower than that in the control group (71%)(P=0.000). (2)There was no statistical difference in cell proliferation between the study group and the control group on day 4, 7, 14 and 21 after CAR-T cell infusion (P=10.223, 3.254, 5.551, 7.605). (3)There was no statistical difference in CRS grading between the study group and the control group (P=0.767). There was no statistical difference in the levels of interleukin 2 receptor (IL-2R), IL-6, tumor necrosis factor (TNF)-α between the two groups. (4)The C-reaction protein (CRP) level of the study group was lower than that of the control group on day 4 and 7 (P=0.026, 0.007). (5)There was no statistical difference of response rates in acute lymphocytic leukemia (ALL) and non-Hodgkin lymphoma (NHL) patients between the two groups (P_ALL=0.842; P_NHL=0.866). Conclusion The modified cell infusion method in CD₁₉ CAR-T cell treatment reduces the incidence of treatment-related FNHTR. It does not affect the proliferation of CAR-T cells in vivo, the grading of CRS and the response rates.

To retrospectively analyze the efficacy and safety of modified cell infusion method in reducing the incidence of febrile non?hemolytic transfusion reaction (FNHTR). Methods A total of 69 patients were enrolled in the clinical trial of CD19 chimeric antigen receptor T (CAR?T) cell treatment from February 2017 to October 2018. Study group received the modified cell infusion method, that 1×106 CAR?T cells were re?suspended in 2 mg human serum albumin with total volume of 20 ml and injected intravenously. The control group was intravenously administrated with CAR?T cell in 100 ml normal saline. The incidence of FNHTR, cytokine releasing syndrome (CRS) grade, cytokine level and efficacy were compared. Results (1)The incidence of FNHTR in the study group was 21.1%, significantly lower than that in the control group (71%)(P=0.000). (2)There was no statistical difference in cell proliferation between the study group and the control group on day 4, 7, 14 and 21 after CAR?T cell infusion (P=10.223, 3.254, 5.551, 7.605). (3)There was no statistical difference in CRS grading between the study group and the control group (P=0.767). There was no statistical difference in the levels of interleukin 2 receptor (IL?2R), IL?6, tumor necrosis factor (TNF)?α between the two groups. (4)The C?reaction protein (CRP) level of the study group was lower than that of the control group on day 4 and 7 (P=0.026, 0.007). (5)There was no statistical difference of response rates in acute lymphocytic leukemia (ALL) and non?Hodgkin lymphoma (NHL) patients between the two groups (PALL=0.842; PNHL=0.866). Conclusion The modified cell infusion method in CD19 CAR?T cell treatment reduces the incidence of treatment?related FNHTR. It does not affect the proliferation of CAR?T cells in vivo, the grading of CRS and the response rates.

Objective: To Evaluation the effect of PD-1 inhibitor Nivolumab on the proliferation and cytotoxicity of anti-CD19 chimeric antigen receptor T cells (CD19-CAR-T) in vitro. Methods: Five patients with high PD-1 expression in peripheral blood and five healthy volunteers were selected. These peripheral blood mononuclear cells were used as the source of T cells to prepare CD19-CAR-T cells. Different doses (72, 36, 18 μg/ml) of Nivolumab was added on day 8 to the culture medium. Patient T cells incubated with 72 μg/ml Nivolumab and CD19-CAR-T cells of healthy volunteers were used as controls. CCK-8, lactate dehydrogenase (LDH) cytotoxicity assay and ELASA were used to detect the proliferation capacity, the specific cytotoxicity and the inflammatory factor secretion. Results: ①T cells from patients with high expression of PD-1 as the source of CD19-CAR-T cells did not affect transfection rate compared with that of healthy volunteers [(32.80±7.22)% vs (35.10±5.84)%, t=-0.554, P=0.593]. ②Incubation of CD19-CAR-T cells with 72 μg/ml Nivolumab did not affect CD19-CAR-T cell proliferation, but its cytotoxicity was significantly higher than that of CD19-CAR-T cells alone or patients’ T cells +72 μg/ml Nivolumab (all P<0.001), there was no significant difference in the killing activity between the 72 μg/ml and 36 μg/ml Nivolumab treated CD19-CAR-T cells on Pfeiffer cells (P=0.281, 0.267, respectively), and they were all higher than those of 18 μg/ml Nivolumab treated CD19-CAR-T cells (all P<0.001). ③Different doses of PD-1 inhibitor Nivolumab combined with CD19-CAR-T cells does not affect the secretion of IFN-γ and IFN-α (all P>0.05). Conclusion Combination of 36 μg/ml PD-1 inhibitor and CD19-CAR-T cells could reduce the drug toxicity and enhance the cytotoxicity.

Objective To analyze the correlation between smoking and occurrence of intracranial artery stenosis.Methods From June 2015 to May 2016,a total of 10 711 inpatients with transient ischemic attack (TIA) or ischemic stroke from 20 basel hospitals of nationwide were enrolled using a cross-sectional study,76 patients with unknown smoking and smoking cessation years were excluded.Finally,a total of 10 635 patients were enrolled.Transcranial color coded sonography and/or transcranial Doppler were used evaluate the intracranial artery stenosis lesions.The basic risk factors for cerebrovascular disease (age,sex,smoking and smoking years,whether smoking cessation and years,hypertension,diabetes,hyperlipidemia,atrial fibrillation,and family history of stroke) were recorded.According to the different smoking years,the smoking years were divided into five groups:non-smoking,smoking time ≤10-year,11 to 20-year,21 to 30-year,and >30-year groups for trend chi square test.According to the different smoking cessation years in the smokers,the smoking cessation years were divided into four groups:non-cessation,cessation time 1 to 10-year,11 to 20-year,and >20-year groups for trend chi square test.The effects of different smoking years and different smoking cessation years on the occurrence of intracranial arterial stenosis were analyzed.Results The incidence of intracranial artery stenosis in the smokers (40.4%[1 433/3 547]) was significantly higher than that in the non-smoking patients (29.4%[2 085/7 088]).There was significant difference (χ2=128.850,P<0.01),and the incidence of cerebral infarction in the smokers (91.6%[3 250/3 547]) was significantly higher than the non-smokers (85.0%[6 027/7 088]).There was significant difference (χ2=92.328,P<0.01).Smoking was an independent risk factor for intracranial artery stenosis (OR,1.603;95%CI 1.456-1.765;P<0.01).With the increase of smoking years,the detection rate of intracranial arterial stenosis increased gradually (trend χ2=115.437,P<0.01).Whether giving up smoking had no significant effect on the incidence of intracranial artery stenosis in patients with ≥20 years of smoking (trend χ2=1.043,P=0.307).Conclusions Smoking is an independent risk factor for affecting intracranial artery stenosis;the risk of disease increases with the number of smoking years.Long-term smokers (≥20 years) cannot reduce the effect on intracranial artery stenosis,even if they give up smoking.