Background and purpose:Chimeric antigen receptor T(CAR-T)cell therapy has shown remarkable efficacy in treating hematological and lymphatic system tumors,but its effectiveness in solid tumors is relatively poor,which is partly attributed to target selection.For Ewing sarcoma(ES),CD99 can be a potential target for CAR-T cells.However,due to T cells'endogenous expression of CD99 protein,CAR-T cells targeting CD99 face limitations in their expansion capacity in vitro.This study aimed to identify the optimal conditions for preparing CD99 CAR-T cells by incorporating CD99 knockdown short hairpin RNA(shRNA),optimizing the multiplicity of infection(MOI)for lentiviral transduction,and screening for the best culture medium and container for CAR-T cell expansion.Methods:shRNA sequences were screened to enhance the expansion capacity of CD99 CAR-T cells.Different MOI,culture media,and containers were used to assess CAR-T cell transduction efficiency,cell viability,proliferation capacity,specific killing ability,and interferon-γ(IFN-γ)release levels under various conditions,in order to identify the optimal cell preparation conditions.Results:The expansion level of KO-CD99 CAR-T cells obtained through shRNA knockdown was significantly higher than that of CD99 CAR-T cells[(16.40±0.40)vs(6.33±1.53),P<0.01].The optimal expansion effect was observed when the transduction MOI was between 0.25 and 1.0,and OptiVitro was used as the culture medium.CAR-T cells cultured in ventilated flasks exhibited significantly higher expansion rates compared to cells cultured in bags[MOI=0.25:(50.23±3.32)vs(13.02±4.82);MOI=0.50:(49.96±0.83)vs(18.25±2.88);MOI=1.00:(48.27±5.08)vs(13.16±6.26);P<0.01],with better cell phenotype and higher specific killing ability.Conclusion:KO-CD99 CAR-T cells obtained through shRNA technology can achieve stable expansion.Based on the optimization of expansion conditions,KO-CD99 CAR-T cells exhibit superior expansion capacity and a higher proportion of memory T cells when the MOI is between 0.25 and 1.00,OptiVitro is used as the culture medium,and ventilated flasks are used as the culture container.These findings lay a solid foundation for further clinical trials of CD99 CAR-T cell therapy for ES.

Objectives:To examine the efficacy of terminal branches portal vein embolization(TBPVE) for the increment of FLR in hepatocellular carcinoma (HCC) patients and to introduce its clinical value with transcatheter chemoembolization(TACE) in the treatment of HCC patients without surgery.Methods:One hundred and fifty HCC patients from three clinical centers of china underwent TBPVE technique from December 2016 to May 2021,including 89 males and 61 females. The average age was 51.9 years(range:18 to 79 years).One hundred and one patients were diagnosed with a background of HBV infection,including 27 patients with portal venous hypertension.TACE was performed simultaneously with TBPVE in 102 patients.Fifty-three patients underwent hepatectomy,who were subdivided into HBV positive and HBV negative groups,with TACE and without TACE groups to analyze the increment of future liver remnant (FLR), complications and survival data.These data were also analyzed in other 97 patients without hepatectomy.Results:All the patients reached adequate FLR successfully in 14 days after TBPVE including patients with portal venous hypertension.The average increment rates of FLR was 56.2% in 7 days and 57.8% in 14 days after TBPVE. There was no significant difference neither between HBV positive and HBV negative groups(7 days:(55.0±27.3)% vs.(57.8±20.9)%, t=0.885, P=0.373; 14 days:(57.3±24.6)% vs.(58.3±23.7)%; t=0.801, P=0.447),or between with TACE and without TACE groups(7 days:(62.3±26.3)% vs. (48.8±20.6)%; t=1.788, P=0.077;14 days:(64.4±25.0)% vs.(55.2±23.1)%; t=1.097, P=0.257).The morbidity and mortality rates were 20.8% and 1.9% in patients with hepatectomy.The 1-,3-year overall survival(OS) and disease-free(DFS) rates were 87.5%,64.5% and 64.7%,40.6% for patients underwent surgery.There was no significant difference of 1-,3-year OS and DFS between HBV positive and negative groups,but there were different between TACE and without TACE groups.The 1-,3-year OS for patients underwent TBPVE and TACE but without surgery were 80.1%, 53.7%. Conclusion:TBPVE is a good alternative technique for modulation of FLR for staged hepatectomy even in HBV positive HCC patients and can be applied with TACE procedure simultaneously as an option treatment for patients with no intend to surgery.

Objectives:To examine the efficacy of terminal branches portal vein embolization(TBPVE) for the increment of FLR in hepatocellular carcinoma (HCC) patients and to introduce its clinical value with transcatheter chemoembolization(TACE) in the treatment of HCC patients without surgery.Methods:One hundred and fifty HCC patients from three clinical centers of china underwent TBPVE technique from December 2016 to May 2021,including 89 males and 61 females. The average age was 51.9 years(range:18 to 79 years).One hundred and one patients were diagnosed with a background of HBV infection,including 27 patients with portal venous hypertension.TACE was performed simultaneously with TBPVE in 102 patients.Fifty-three patients underwent hepatectomy,who were subdivided into HBV positive and HBV negative groups,with TACE and without TACE groups to analyze the increment of future liver remnant (FLR), complications and survival data.These data were also analyzed in other 97 patients without hepatectomy.Results:All the patients reached adequate FLR successfully in 14 days after TBPVE including patients with portal venous hypertension.The average increment rates of FLR was 56.2% in 7 days and 57.8% in 14 days after TBPVE. There was no significant difference neither between HBV positive and HBV negative groups(7 days:(55.0±27.3)% vs.(57.8±20.9)%, t=0.885, P=0.373; 14 days:(57.3±24.6)% vs.(58.3±23.7)%; t=0.801, P=0.447),or between with TACE and without TACE groups(7 days:(62.3±26.3)% vs. (48.8±20.6)%; t=1.788, P=0.077;14 days:(64.4±25.0)% vs.(55.2±23.1)%; t=1.097, P=0.257).The morbidity and mortality rates were 20.8% and 1.9% in patients with hepatectomy.The 1-,3-year overall survival(OS) and disease-free(DFS) rates were 87.5%,64.5% and 64.7%,40.6% for patients underwent surgery.There was no significant difference of 1-,3-year OS and DFS between HBV positive and negative groups,but there were different between TACE and without TACE groups.The 1-,3-year OS for patients underwent TBPVE and TACE but without surgery were 80.1%, 53.7%. Conclusion:TBPVE is a good alternative technique for modulation of FLR for staged hepatectomy even in HBV positive HCC patients and can be applied with TACE procedure simultaneously as an option treatment for patients with no intend to surgery.

Objective To investigate the efficacy and safety of percutanous transhepatic intrahepatic portosystemic shunt(PTIPS)for chronic portal vein occlusion and cavernous transformation with symptomatic portal hypertension.Methods The clinical and imaging data of 38 patients with chronic portal vein occlusion and cavernous transformation with symptomatic portal hypertension, who received PTIPS in our hospital from November 2009 to June 2016,were analyzed retrospectively.The differences of the portosystemic pressure gradient(PPG)measured before and after PTIPS procedure was analyzed by a paired samples t-test. All the patients were followed up and the curative effect and operation-correlated complications were observed.Results The PTIPS procedure was technically successful in 36 patients.The other two patients with unsuccessful PTIPS underwent medical treatment,and one of them died of recurrent variceal bleeding 25 months later. Effective portal decompression and free antegrade shunt flow were achieved in 36 patients with successful PTIPS.And the mean PPG was decreased from(25.2±2.9)to(13.2± 1.3) mmHg (1 mmHg=0.133 kPa) before and after PTIPS respectively and the difference was statistically significant(P<0.05).During the procedure,arterial hemorrhage occurred in two patients who subsequently underwent embolization. Biliary injury occurred in one case and percutanous transhepatic biliary drainage (PTBD)was then performed.The mean follow-up period of the 36 patients was(26.7±10.4)months(range from 3.0 to 74.0 months).Hepatic encephalopathy appeared in 4 cases,among which,3 patients recovered after receiving medical treatment, while 1 patient experienced Grade 3 hepatic encephalopathy and recovered after implanting a smaller cover-stent.Shunt dysfunction occurred in 10 cases,of which 8 cases recovered after shunt revision with stent implantation or ballon angioplasty, while 2 cases underwent anticoagulation by warfarin only. During follow-up period, 7 patients died of liver failure(n=4), hepatic cellular carcinoma(n=1), recurrent varicose vein bleeding(n=1), and renal failure(n=1). The other patients remained asymptomatic and shunt patency. Conclusions PTIPS is both safe and effective for the treatment of symptomatic portal hypertension caused by chronic portal vein occlusion and cavernous transformation.The technical success rate is high,and the short-term curative effect is satisfied.

Objective To compare the anesthetic effect of ropivacaine and bupivacaine in combined spinal-epidural anesthesia(CSEA)for cesarean section,and their influence on the incidence rate of supine hypotension syndrome(SHS).Methods 200 patients with cesarean section surgery in our hospital from February 2016 to July 2016 were randomly divided into observation group and control group,all of cases were given CSEA.100 patients in the observation group(the group L)were given ropivacaine in spinal anesthesia,the other 100 patients in the control group(the group B)were given bupivacane in spinal anesthesia.Recorded the relevant indicators,compared the incidence rate of SHS,the effect of anesthesia and neonatal score.Results The incidence rate of SHS of the group L was lower than the group B(χ2 =9.261,P<0.01).The effect of anesthesia and Apgar score of two groups had no statistically significant differences(all P>0.05).Conclusion The application of ropivacaine in CSEA for cesarean section not only has exact anesthesia effect,but also can effectively prevent SHS without any side effects.

Objective: To analyze the efficacy of branches portal vein embolization (TBPVE) combined with transcatheter arterial chemoembolization (TACE) on liver neoplasms. Methods: From August 2016 to May 2017, there were 13 patients including 11 males and 2 females with primary hepatocellular carcinoma who underwent TBPVE+ TACE , among whom there were 11 cases with a history of HBV infection.Average age of the 13 patients was (60.8±6.2)years. The live function of all patients were Child-Pugh A classification.The CT or MRI images of each patient was reconstructed and the standard liver volume(SLV) before TBPVE+ TACE was (1 181.2±49.3)ml, estimated future liver remnant(FLR) was (326.1±72.1)ml and FLR/SLV was (27.6±6.0)%.The puncture site for TBPVE was determined by the three-dimensional reconstruction of portal vein.CT scan or MRI, AFP and liver function test were repeated after one and two weeks after TBPVE+ TACE.FLR and FLR/SLV were calculated respectively.Hepatectomy would be performed if the patients agreed.The postoperative complications were analyzed. Results: On the 7^thday after TBPVE+ TACE, the FLR/SLV was(42.6±8.0)% and the FLR increasement was(56.0±24.6)%.The level of AFP decreased from(87.9±81.8)μg/L to (29.7±20.9)μg/L.On the 14^thday after TBPVE+ TACE, the FLR/SLV was(45.8±6.2)% and the FLR increasement was(71.8±29.0)%.Four patients underwent surgery which including 2 right hepatectomies and 2 right trisegmentectomies in 2 weeks after TBPVE+ TACE.Nine patients were performed with targeting intratumoral lactic acidosis TACE (TILA-TACE). No severe complication occurred in all patients. Conclusions TBPVE could induce a rapid growth of the liver remnant but still with the concern of inducing the growth of neoplasms at the same time.To combine TACE in TBPVE therapy not also can the growth of neoplasms be prevented but also inducing its shrinking.This method might be a new mode for the treatment of hepatocellular carcinoma.

Objective To investigate the risk factors of cerebral microbleeds (CMBs) in patients with acute isch?emic stroke of large-artery atherosclerosis. Methods One hundred twelve patients with acute ischemic stroke of large-ar?tery atherosclerosis admitted from July 2013 to January 2014 in Nanjing First Hospital affiliated to Nanjing Medical Uni?versity were enrolled. According to the results of MRI magnetic sensitive weighted imaging, the patients were divided into CMBs group or non-CMBs group. The history, general clinical data, serum biochemical results and MRI in both groups were enrolled. All the data were analyzed by the univariate and multivariate analysis. Results The results of univariate analysis showed that there were significant differences in age (61.620±11.479 vs. 70.620±11.185), serum uric acid (UA) level (278.920±69.512 vs. 353.460±111.206), serum creatinine (Cr) level (71.360±19.797 vs. 90.450±44.989), serum ho?mocysteine (Hcy) level (12.587±2.664 vs. 21.715±10.437) between the two groups (P<0.05). There were significant differ?ences in constituent ratio of Fazekas' s grade of periventricular hyperintensities and deep white matter hyperintensities between the two groups (P<0.05). The results of multivariate analysis showed that age (OR=0.963, 95%CI:0.905~1.025, P<0.05) and serum Hcy level (OR=1.487, 95%CI:1.219~1.813, P<0.05) were the independent risk factors for CMBs in patients with acute ischemic stroke of large-artery atherosclerosis. Conclusions Age and serum Hcy level are the inde?pendent risk factors for CMBs in patients with acute ischemic stroke of large-artery atherosclerosis.

Objective To investigate the safety and efficacy of interventional treatment for ischemic-type biliary lesion (ITBL)after liver transplantation (LT). Methods The clinical and imaging data of 76 patients with ITBL after LT,who received interventional treatment in the Department of Interventional Vascular Radiology of the Third Affiliated Hospital of Sun Yat-sen University from January 2006 to February 2014,were retrospectively analyzed. On the basis of the cholangiographic appearance,patients were classified into 3 groups:hilar biliary stricture group (n=28),multifocal biliary stricture group (n=42),and biloma group (n=6). The modalities of interventional treatment were percutanous transhepatic biliary drainage (PTBD), PTBD combined with balloon dilation,PTBD combined with balloon dilation and plastic stent implantation. The methods of biliary drainage included external drainage and external-internal drainage. All the patients were followed up after treatment. The curative effect and biliary complication was observed. Results The first successful rate of PTBD was 97% (74/76). The total curative rate,improvement rate and ineffective rate of interventional treatment were 21% (16/76),51% (39/76)and 28% (21/76). In hilar biliary stricture group,the cure,improvement and inefficacy rates were 36% (10/28),57% (16/28)and 7% (2/28).The efficacy rate was 93% (22/28 ). In multifocal biliary stricture group,the cure,improvement and inefficacy rates were 14% (6/42),50% (21/42)and 36% (15/42). The efficacy rate was 64% (27/42). In biloma group,2 cases (2/6)were cured and treatment of4 cases was ineffective. The efficacy ofhilar biliary stricture group was better than that of multifocal biliary stricture group (P<0.05 ). The efficacy of multifocal biliary stricture group was better than that of biloma group (P <0.001 ). The main biliary complication was biliary tract infection during drainage. The rates of bile tract infections were 20% (13/64) and 67% (8/12)in patients with external drainage and external-internal drainage,respectively. There was significant difference between these two items (P <0.001 ). Conclusions PTBD is a safe and effective therapeutic modality for ITBL after LT,which combined with balloon dilation and biliary stent implantation can improve patients’clinical symptoms,elevate patients’quality of life. The biliary external drainage can decrease the rate of biliary tract infection significantly.

OBJECTIVETo investigate the evaluation of lymphatic emboli by D2-40/CKpan dual immunostain and its prognostic significance in advanced primary gastric adenocarcinoma.

METHODSTissue samples of 108 cases of advanced gastric adenocarcinoma with radical gastrectomy were collected from Chinese PLA General Hospital in 2001. Lymphatic emboli were evaluated by D2-40/CKpan dual immunostain on consecutive sections, and compared with routine HE staining. The correlation of lymphatic emboli with lymph node metastasis and overall survival was analyzed by Pearson Chi-squared test and univariate survival analysis, respectively.

RESULTSLymphatic emboli were detected in 73/108 (67.6%) cases by D2-40/CKpan staining, compared to 57/108 (52.8%) by HE staining. There was significant difference (P=0.007). Lymphatic emboli on HE staining revealed false negative in 24/108 cases and false positive in 8/108 cases. A significant correlation was found between lymphatic emboli and lymph node metastasis on HE staining (P=0.024), while it was not found on D2-40/CKpan staining (P=0.422). The overall survival rate was significantly different between lymphatic emboli positive and negative cases on HE staining (P=0.043). The overall survival rate was lower in lymphatic emboli positive cases on HE staining. Lymphatic emboli evaluated on D2-40/CKpan staining had no prognostic value (P=0.402).

CONCLUSIOND2-40/Ckpan dual immunostaining is more sensitive for lymphatic emboli in gastric adenocarcinoma but may not predict lymph node metastasis and survival, while HE staining may.

Objective To investigate the effect of candesartan cilexetil on rotenone-induced Parkinson's disease (PD) in rats.Methods Forty 10-week-old male Lewis rats were chosen in our study and equally randomized into control group,rotenone group,rotenone+candesartan cilexetil group and candesartan cilexetil group (n=10); rotenone (2.5-3.0 mg/[kg· d]) was given for 4 weeks to the rats of rotenone group and rotenone+candesartan cilexetil group by subcutaneous osmotic minipumps implantation under the back; rats in the rotenone+candesartan cilexetil group and candesartan cilexetil group were orally administered candesartan cilexetil.Neurological behavioral measurements were performed to evaluate the motor features; tyrosine hydroxylase (TH) and α-synuclein immunoreactivities in the substantia nigra pars compacta (SNc) were observed.Protein level of α-synuclein was determined by Westem blotting.Results The weight of rats in the rotenone group reduced to (297.3±12.2) g,with significant difference as compared with that of the other three groups (P＜0.05); decreased TH immunoreactivity (377.0±41.6) cells/mm2) and increased α-synuclein immunoreactivity (0.75±0.02) in the SNc of rats in the rotenone group was noted,enjoying significant differences as compared with the other three groups (P＜0.05); these values in the rotenone+candesartan cilexetil group were (337.2±26.3) g,(639.7±46.0) cells/mm2 and 0.57±0.01,respectively (P＜0.05).Western blotting confirmed that rotenone up-regulated the expression ofα-synuclein in the SNc,and candesartan ceilexetil markedly attenuated the increase (P＜0.05).Conclusion Candesartan cilexetil can protect rotenone-induced PD in rats through decreasing TH-positive cell apoptosis and α-synuclein deposition.