Pancreatic cancer has a very poor prognosis.Early diagnosis and treatment are especially critical for improving its prognosis.Nanotechnology has been widely used in the diagnosis and treatment of pancreatic cancer.Relying on the unique physicochemical properties of nanoparticles and their rich surface modifications,effective enrichment of tumor sites can be achieved.Magnetic iron oxide nanoparticles(MIONPs)is one of the commonly used nanomaterials in the diagnosis and treatment of pancreatic cancer,and has good biocompatibility.Through special surface modification,it can be used in targeted diagnosis and treatment of pancreatic cancer.MIONPs can be used as a contrast agent for MRI,and by modifying the surface,they also can be used in targeted imaging of pancreatic cancer.And they can also be modified as a drug delivery system to achieve targeted delivery of drugs and improve therapeutic effects.However,the application of MIONPs in pancreatic cancer diagnosis and treatment still faces some challenges,such as nanotoxicity and cost issues.With the development of technology,MIONPs are expected to play an important role in the personalized diagnosis and treatment of pancreatic cancer.

Objective:To investigate the effect of copper chlorophyllin sodium salt(CHL)on the sensitivity of human pancreatic cancer cells in response to gemcitabine(GEM)therapy and on the therapeutic effect on pancreatic cancer cells that have developed GEM resistance.Methods:MIA GR(a pancreatic cancer cell line resistant to GEM)was induced by a low-dose continuous incremental method,and the half inhibitory concentration(IC50)of MIA WT and MIA GR to GEM treatment was detected by the CCK-8 method,and the resistance index was calculated;the difference in IC50 of CHL on the two types of cells was detected by the CCK-8 method after treating MIA WT and MIA GR cells with different concentrations of CHL,CCK-8 method was used to detect the difference in IC50 of CHL on the two types of cells;on the basis of IC50,MIA WT and MIA GR cells were intervened with CHL and(or)GEM with different multiplicity of IC50,respectively,and the growth inhibition curves of MIA WT and MIA GR cells were detected by the CCK-8 method under the intervention of CHL combined with GEM;After the intervention of MIA WT and MIA GR cells with CHL and(or)GEM at IC50,respectively,the effects on the proliferation of the two different cells were detected using the clone formation assay;the effects on cytotoxicity/activity were observed under fluorescence microscopy;and the effects on apoptosis were detected using flow cytometry.Finally,western blotting was used to detect the effects of CHL and(or)GEM interventions on the drug resistance-associated molecules P-glycoprotein(P-gp)and ribonucleotide reductase regulatory subunit M2(RRM2)in MIA GR cells,the and sensitivity-related molecule deoxycytidine kinase(DCK)on protein expression levels.Results:MIA GR cells were verified to be well drug resistant,with resistance indices of 549.1 and 667.9 after 48 h and 96 h after GEM intervention compared to homologous wild-type MIA WT cells,respectively;CHL intervention inhibited the proliferation of MIA GR cells more significantly compared to that of MIA WT cells;and CHL in combination with GEM exerted a more significant growth inhibitory effect compared to GEM alone in both MIA WT cells(P<0.001)and MIA GR cells(P<0.01).CHL significantly inhibited the tumor proliferation of MIA GR cells,and the inhibitory effect was more pronounced in both cells when combined with GEM(P<0.000 1);furthermore,compared to GEM alone,the intervention with CHL could cause more pronounced cytotoxicity(P<0.000 1)in both MIA WT and MIA GR cells.caused more pronounced cytotoxicity(P<0.000 1)and induced a higher percentage of apoptosis than GEM alone.The results of the western blotting assay showed that CHL intervention caused a decrease in the expression levels of P-gp and RRM2 proteins,as well as an increase in the protein expression level of DCK in MIA GR cells.Conclusion:CHL increases the sensitivity of pancreatic cancer cells to GEM and also induces a decrease in the resistance of drug-resistant pancreatic cancer cells to GEM.

Objective:To investigate the effect of copper chlorophyllin sodium salt(CHL)on the sensitivity of human pancreatic cancer cells in response to gemcitabine(GEM)therapy and on the therapeutic effect on pancreatic cancer cells that have developed GEM resistance.Methods:MIA GR(a pancreatic cancer cell line resistant to GEM)was induced by a low-dose continuous incremental method,and the half inhibitory concentration(IC50)of MIA WT and MIA GR to GEM treatment was detected by the CCK-8 method,and the resistance index was calculated;the difference in IC50 of CHL on the two types of cells was detected by the CCK-8 method after treating MIA WT and MIA GR cells with different concentrations of CHL,CCK-8 method was used to detect the difference in IC50 of CHL on the two types of cells;on the basis of IC50,MIA WT and MIA GR cells were intervened with CHL and(or)GEM with different multiplicity of IC50,respectively,and the growth inhibition curves of MIA WT and MIA GR cells were detected by the CCK-8 method under the intervention of CHL combined with GEM;After the intervention of MIA WT and MIA GR cells with CHL and(or)GEM at IC50,respectively,the effects on the proliferation of the two different cells were detected using the clone formation assay;the effects on cytotoxicity/activity were observed under fluorescence microscopy;and the effects on apoptosis were detected using flow cytometry.Finally,western blotting was used to detect the effects of CHL and(or)GEM interventions on the drug resistance-associated molecules P-glycoprotein(P-gp)and ribonucleotide reductase regulatory subunit M2(RRM2)in MIA GR cells,the and sensitivity-related molecule deoxycytidine kinase(DCK)on protein expression levels.Results:MIA GR cells were verified to be well drug resistant,with resistance indices of 549.1 and 667.9 after 48 h and 96 h after GEM intervention compared to homologous wild-type MIA WT cells,respectively;CHL intervention inhibited the proliferation of MIA GR cells more significantly compared to that of MIA WT cells;and CHL in combination with GEM exerted a more significant growth inhibitory effect compared to GEM alone in both MIA WT cells(P<0.001)and MIA GR cells(P<0.01).CHL significantly inhibited the tumor proliferation of MIA GR cells,and the inhibitory effect was more pronounced in both cells when combined with GEM(P<0.000 1);furthermore,compared to GEM alone,the intervention with CHL could cause more pronounced cytotoxicity(P<0.000 1)in both MIA WT and MIA GR cells.caused more pronounced cytotoxicity(P<0.000 1)and induced a higher percentage of apoptosis than GEM alone.The results of the western blotting assay showed that CHL intervention caused a decrease in the expression levels of P-gp and RRM2 proteins,as well as an increase in the protein expression level of DCK in MIA GR cells.Conclusion:CHL increases the sensitivity of pancreatic cancer cells to GEM and also induces a decrease in the resistance of drug-resistant pancreatic cancer cells to GEM.

The chronic use of morphine and other opioids is associated with opioid-induced hypersensitivity (OIH) and analgesic tolerance. Among the different forms of OIH and tolerance, the opioid receptors and cell types mediating opioid-induced mechanical allodynia and anti-allodynic tolerance remain unresolved. Here we demonstrated that the loss of peripheral μ-opioid receptors (MORs) or MOR-expressing neurons attenuated thermal tolerance, but did not affect the expression and maintenance of morphine-induced mechanical allodynia and anti-allodynic tolerance. To confirm this result, we made dorsal root ganglia-dorsal roots-sagittal spinal cord slice preparations and recorded low-threshold Aβ-fiber stimulation-evoked inputs and outputs in superficial dorsal horn neurons. Consistent with the behavioral results, peripheral MOR loss did not prevent the opening of Aβ mechanical allodynia pathways in the spinal dorsal horn. Therefore, the peripheral MOR signaling pathway may not be an optimal target for preventing mechanical OIH and analgesic tolerance. Future studies should focus more on central mechanisms.
Humans ; Morphine/pharmacology* ; Hyperalgesia/metabolism* ; Analgesics, Opioid/pharmacology* ; Neurons/metabolism* ; Signal Transduction

Objective To investigate the guiding role of quantitative evaluation system of immune status in the individualized management of immunosuppressants for the recipients after liver transplantation. Methods Clinical data of 239 liver transplant recipients were retrospectively analyzed. MingDao Immune Cell Analysis (MICA) was established. All recipients were divided into two groups according to the adjustment regimens of immunosuppressants. The immunosuppressant regimen was adjusted according to MingDao Immune System Score (MISS) in the MISS group (n=84), and the medication plan was empirically adjusted during the same period in the control group (n=155). According to the time of postoperative detection (t), the recipients in the MISS group were divided into subgroup A (t ≤ 28 d, n=78), subgroup B (28 d < t ≤ 6 months, n=68), subgroup C (6 months < t ≤ 12 months, n=18), subgroup D (12 months < t ≤ 24 months, n=18) and subgroup E (t > 24 months, n=19). In the MISS group, postoperative MISS scores of recipients in subgroups A-E were analyzed. The incidence of acute rejection and opportunistic infection and the overall survival rate were statistically compared between the MISS and control groups. Results The MISS scores in subgroups A-E were -7.0 (-13.2, -2.0), -2.0 (-5.8, 1.8), -0.5 (-7.3, 2.8), -2.0 (-4.5, 3.3) and -3.0 (-6.0, 1.0), respectively. The immune status of the recipients was gradually improved over postoperative time, and the difference between two groups was statistically significant (P < 0.05). In the MISS group, 15% (13/84) of the recipients developed acute rejection, and 27% (42/155) in the control group, and the difference was statistically significant (P < 0.05). In the MISS group, the MISS score of the recipients with acute rejection was 0 (-2.5, 3.5), and -5.0 (-12.0, -1.0) for their counterparts without acute rejection, and the difference was statistically significant (P < 0.05). In the MISS group, 2% (2/84) of the recipients presented with postoperative opportunistic infection, and 9% (14/155) in the control group, and the difference was statistically significant (P < 0.05). In the MISS group, the 1- and 3-year overall survival rates were 86.9% and 79.8%, and 83.2% and 76.8% in the control group, and no significant difference was observed between two groups (P > 0.05). Conclusions MICA and MISS score may reflect the immune status of liver transplant recipients, and guide the individualized management of administration of immunosuppressants after liver transplantation.

[摘 要] 目的：探讨miR-1243通过靶向调控核不均一核糖核蛋白A2/B1（hnRNPA2B1）表达对肝癌HepG2细胞增殖、迁移的影响及其分子机制。方法：用qPCR和WB法检测40例肝癌组织及其癌旁组织（2019年1月至2021年8月在武汉市第三医院首义院区手术切除标本）和正常人肝细胞QSG-7701与肝癌细胞HepG2、Hep3b、HuH-7中miR-1243、hnRNPA2B1 mRNA水平及hnRNPA2B1、cyclin D1、MMP-2蛋白水平；双荧光素酶报告基因实验验证miR-1243和hnRNPA2B1的靶向关系。HepG2细胞分为对照组（不转染）、miR-NC组（转染miR-NC）、miR-1243 mimic组（转染miR-1243 mimic）、miR-1243 mimic+pcDNA3.1组（转染miR-1243 mimic和pcDNA3.1）、miR-1243 mimic+pc-hnRNPA2B1组（转染miR-1243 mimic和pc-hnRNPA2B1）后进行相应转染；WB法检测肝癌组织及细胞和转染后各组细胞的hnRNPA2B1、cyclin D1、MMP-2蛋白表达水平；CCK-8法检测转染后各组HepG2细胞的增殖能力；划痕愈合实验检测转染后各组HepG2细胞的迁移能力。结果：与癌旁组织或正常人肝细胞QSG-7701相比，肝癌组织和肝癌细胞中miR-1243呈低表达、hnRNPA2B1 mRNA及其蛋白呈高表达（均P<0.05）。双荧光素酶报告基因实验结果证实miR-1243与hnRNPA2B1间存在靶向关系，且miR-1243通过靶向hnRNPA2B1负调控其表达。转染miR-1243 mimic后HepG2细胞中hnRNPA2B1蛋白表达、细胞增殖能力、划痕愈合率及cyclin D1、MMP-2蛋白表达均显著降低（均P<0.05）；而同时过表达hnRNPA2B1和miR-1243可逆转过表达miR-1243对HepG2细胞增殖、迁移的抑制作用。结论：miR-1243通过靶向hnRNPA2B1表达调控肝癌HepG2细胞的增殖和迁移。

[Abstract] Objective: To investigate the effects of long non-coding RNA zinc finger antisense 1 (lncRNA ZFAS1) on the proliferation, invasion, and migration of liver cancer cells by regulating the miR-588/high mobility group AT-hook protein 2 (HMGA2) axis. Methods: Real-time fluorescent quantitative PCR (qRT-PCR) and western blot were performed to measure the expression levels of ZFAS1, miR-588 and HMGA2 in 80 pairs of liver cancer tissues and corresponding para-cancerous issues (the tissue sample were collected from Shouyi Campus, Wuhan Third Hospital during Jan. 2018 and Dec. 2019), human normal liver cell line (LO2) and liver cancer cell lines (HepG2, Huh7, HCCLM3). The survival of patients was analyzed using the Kaplan-Meier survival curve. HepG2 cells were divided into blank group, si-NC group, si-ZFAS1 group, si-ZFAS1+inhibitor NC group, and si-ZFAS1+miR-588 inhibitor group. qPCR was performed to measure the expression of ZFAS1 and miR-588 in HepG2 cells of each group, and Western blot was performed to measure the expression of HMGA2 protein in the cells. CCK-8 method, Transwell, and scratch test were performed to measure the proliferation, invasion, and migration of HepG2 cells. Dual-luciferase reporter gene experiment was performed to verify the targeting relationship between ZFAS1 and miR-588 as well as between miR-588 and HMGA2. A HepG2 cell transplanted tumor model was established in nude mice to examine the effect of silencing ZFAS1 or/and miR-588 on the growth of transplanted tumors. Results: ZFAS1 and HMGA2 were highly expressed while miR-588 was lowly expressed in liver cancer tissues and liver cancer cells (all P<0.05). The 2-year survival rate of patients with low ZFAS1 expression was higher than that in the high expression group (P<0.05). Compared with the blank group, the relative expression of ZFAS1 and HMGA2 protein in the si-ZFAS1 group was significantly reduced, and the relative expression of miR-588 was significantly increased (all P<0.05); compared with the si-ZFAS1 group, the relative expression of ZFAS1 in the si-ZFAS1+miR-588 inhibitor group did not change significantly (P>0.05), however, the relative expression of HMGA2 protein was significantly increased, and the relative expression of miR-588 was significantly reduced (P<0.05). Silencing ZFAS1 was able to inhibit the proliferation, invasion, and migration of HepG2 cells and inhibit the growth of transplanted tumors in nude mice (all P<0.05). ZFAS1 targeted and down-regulated the expression of miR-588, while miR-588 targeted and down-regulated the expression of HMGA2. Simultaneous inhibition of miR-588 expression could reverse the inhibitory effects of silencing ZFAS1 on the proliferation, invasion, and migration of HepG2 cells and the growth of transplanted tumors in nude mice (all P<0.05). Conclusion: Silencing ZFAS1 may down-regulate the expression of HMGA2 by promoting the expression of miR-588, thereby inhibiting the proliferation, invasion, and migration of liver cancer HepG2 cells.

Objective:We proposed a Mingdao immune score system(MISS)to evaluate recipient's immune status after liver transplantation.Methods:From January 2017 to June 2019, retrospective analysis was conducted for 89 recipients of liver transplantation. Age/gender-matched 385 healthy controls(HC)were selected. The percentages of 30 lymphocyte subgroups of patients and HC were measured by flow cytometry. The score of each individual was calculated with our proposed MISS method. And drug concentrations and relevant clinical data were collected.Results:The normal MISS value of a healthy person was 0 score according to our criterion. In this study, the value of MISS for HC was distributed in a nearly normal fashion(-0.73±4.02). When the data from patients at different timepoints were compared, the MISS value started with -1.21±7.42 pre-operation, then declined sharply down to -8.95±8.05 at 1 month and jumped to -4.50±7.80 at 3 months. Afterward it stabilized at -4.18±7.83 between 3~12 months post-operation and finally reached -2.00±5.51 at 1 year ( P<0.05). Patients with acute rejection had higher MISS values than those without acute rejection, ( P<0.05). No significant correlation existed between blood drug concentrations and MISS values ( P>0.05). Conclusions:Our proposed MISS method may reflect the whole immune status. It is useful to manage the application of immunosuppressants in conjunctions with blood drug concentrations and liver graft function.

Objective To explore the clinical efficacy of liver transplantation for severe liver disease. Methods The clinical data of 51 patients who underwent liver transplantation for severe liver disease were retrospectively analyzed. The general intraoperative conditions were observed, including operation duration, warm ischemia time, cold ischemia time, anhepatic phase, bleeding volume, blood transfusion volume, plasma transfusion volume and so on. The changes in indexes such as total bilirubin (TB), prothrombin time activity (PTA), and prothrombin time international normalized ratio (PT-INR) were observed before operation and at 3 d, 1 week and 2 weeks after operation. The postoperative survival and occurrence of complications were analyzed. The indexes that might affect the prognosis of patients with severe liver disease were analyzed by Cox regression analysis. Results For the 51 patients, operation duration, warm ischemia time and cold ischemia time was 8 (7, 9) h, 3 (2, 3) min and 6 (5, 8) h respectively, intraoperative anhepatic phase was 80 (70, 100) min, intraoperative bleeding volume was 1 000 (550, 1 500) mL, and intraoperative blood transfusion volume was 1 200 (200, 1 600) mL. Postoperative TB, PTA, PT-INR and other indexes improved significantly compared to those preoperatively. Among the 51 patients, 10 cases died, with the death causes of multiple organ failure and severe infection(7 cases), renal insufficiency (2 cases), and cardiovascular complications (1 case). Survival rates at 1 month and 1 year post-transplantation for patients with severe liver disease were 82% and 80%, respectively. Cox regression analysis suggested that abnormal preoperative PTA and PT-INR were the risk factors for post-transplantation death in patients with severe liver disease. Conclusions Liver transplantation significantly improves the survival rate for patients with severe liver disease, perioperative infection prevention and treatment as well as multiple organ function management play key roles in improving post-transplantation survival rate in patients with severe liver disease.

Objective To investigate the effect of Immutol on inducing the immune tolerance of cardiac grafts in rat models. Methods A rat model of heterotopic abdominal heart transplantation was established. The recipient rats were divided into 5 groups: blank control group (n=6); dimethyl sulfoxide (DMSO) group (n=6), in which DMSO was administered until the cardiac graft arrest; Immutol group (n=6), in which Immutol was administered until the cardiac graft arrest; ciclosporin (CsA) group (n=10), in which CsA was administered for 20 d; combined group (n=13), in which Immutol was given for 60 d combined with CsA for 20 d. The survival time and pathological changes of cardiac grafts in each group were observed. The contents of serum interleukin (IL)-10 and interferon (IFN)-γ were detected. The expression levels of indoleamine 2, 3-dioxygenase (IDO) and fibrinogen-like protein 2(Fgl2) messenger RNA(mRNA) in heart tissues of rats in each group were measured. Results In the combined group, the cardiac grafts survived for >180 d and immune tolerance was induced. The pathological score of cardiac grafts in the combined group was significantly lower than that in the CsA group at postoperative 39 d (P < 0.05). The levels of serum IL-10 and IFN-γ in the combined group were significantly higher than those in the CsA group at 9 d and 39 d after operation (both P < 0.05). The content of serum IL-10 and IFN-γ in the combined group were gradually increased over time. At postoperative 39 d, the expression levels of IDO and Fgl2 mRNA in the combined group were significantly higher than those in the CsA group (both P < 0.05). The expression level of IDO mRNA in the combined group tended to gradually elevate after operation. In the combined group, the expression level of Fgl2 mRNA at postoperative 180 d was significantly higher than those at 9 d and 39 d after operation (both P < 0.05). Conclusions Combined administration of Immutol and CsA can effectively inhibit the incidence of acute rejection, and maintain the long-term survival of the cardiac grafts and induce immune tolerance after drug withdrawal.