OBJECTIVE: To explore the construction method of a resistant multiple myeloma (MM) patient-derived xenotransplantation (PDX) model. METHODS: 1.0×10⁷ MM patient-derived mononuclear cells (MNCs), 2.0×10⁶ MM.1S cells and 2.0×10⁶ NCI-H929 cells were respectively subcutaneously inoculated into NOD.CB17-Prkdc^scid Il2rg^tm1/Bcgen (B-NDG) mice with a volume of 100 μl per mouse to establish mouse model. The morphologic, phenotypic, proliferative and genetic characteristics of PDX tumor were studied by hematoxylin-eosin staining, immunohistochemical staining (IHC), cell cycle analysis, flow cytometry and fluorescence in situ hybridization (FISH). The sensitivity of PDX tumor to bortezomib and anlotinib monotherapy or in combination was investigated through cell proliferation, apoptosis and in vitro and in vivo experiments. The effects of anlotinib therapy on tumor blood vessel and cell apoptosis were analyzed by IHC, TUNEL staining and confocal fluorescence microscope. RESULTS: MM PDX model was successfully established by subcutaneously inoculating primary MNCs. The morphologic features of tumor cells from MM PDX model were similar to those of mature plasma cells. MM PDX tumor cells positively expressed CD138 and CD38, which presented 1q21 amplification, deletion of Rb1 and IgH rearrangement, and had a lower proliferative activity than MM cell lines. in vitro, PDX, MM.1S and NCI-H929 cells were treated by bortezomib and anlotinib for 24 hours, respectively. Cell viability assay showed that the IC₅₀ value of bortezomib were 5 716.486, 1.025 and 2.775 nmol/L, and IC₅₀ value of anlotinib were 5 5107.337, 0.706 and 5.13 μmol/L, respectively. Anlotinib treatment increased the apoptosis of MM.1S cells (P < 0.01), but did not affect PDX tumor cells (P >0.05). in vivo, there was no significant difference in PDX tumor growth between bortezomib monotherapy group and control group (P >0.05), while both anlotinib monotherapy and anlotinib combined with bortezomib effectively inhibited PDX tumor growth (both P < 0.05). The vascular perfusion and vascular density of PDX tumor were decreased in anlotinib treatment group (both P < 0.01). The apoptotic cells in anlotinib treatment group were increased compared with those in control group (P < 0.05). CONCLUSION Bortezomib-resistant MM PDX model can be successfully established by subcutaneous inoculation of MNCs from MM patients in B-NDG mice. This PDX model, which retains the basic biological characteristics of MM cells, can be used to study the novel therapies.
Animals ; Bortezomib ; Humans ; Multiple Myeloma/pathology* ; Mice ; Apoptosis ; Drug Resistance, Neoplasm ; Cell Line, Tumor ; Xenograft Model Antitumor Assays ; Mice, Inbred NOD ; Disease Models, Animal ; Cell Proliferation ; Transplantation, Heterologous

OBJECTIVE: This study aimed to determine the expression levels and prognostic significance of MYCN in bone marrow of adult patients with newly diagnosed acute myeloid leukemia (AML). METHODS: A total of 62 newly diagnosed patients with non-M3 AML were enrolled as the study group, and 20 healthy donors as the control group. Real-time quantitative reverse transcription-polymerase chain reaction (PCR) was performed to detect the expression level of MYCN, and the relationship between MYCN expression and prognosis of AML patients was analyzed. RESULTS: MYCN was up-regulated in newly diagnosed AML patients compared with normal controls (P < 0.001). Receiver operating characteristic (ROC) curve analysis revealed that MYCN could serve as a diagnostic biomarker for AML. Kaplan-Meier survival analysis showed that the patients with high MYCN expression had a shorter overall survival (OS) time than the patients with low MYCN expression (P =0.016). The expression level of MYCN was lower during the complete ressimion (CR) phase of AML compared to the initial diagnosis, but it returned to the initial diagnostic level or even higher during relapse phase. Multivariate Cox regression analysis showed that high expression of MYCN was an independent risk factor for OS of AML patients (P =0.021). CONCLUSION MYCN is highly expressed and associated with poor prognosis in de novo AML, which might be serve as a novel diagnostic and prognostic biomarker for adult AML.
Humans ; Leukemia, Myeloid, Acute/metabolism* ; Prognosis ; N-Myc Proto-Oncogene Protein ; Adult ; Female ; Male ; Middle Aged

Objective:To analyze the efficacy and feasibility of performing a new surgical procedure, tunnel esophagogastrostomy, to perform proximal gastrectomy.Methods:The study cohort comprised 10 consecutive patients who had undergone esophagogastrostomy by the tunnel technique in Jiangsu Cancer Hospital between October 2019 and July 2022. All patients were male. Their average age was (64.2±8.1) years and body mass index (25.5±3.2) kg/m2. Nine had upper gastric body adenocarcinoma, the remaining one having signet ring cell carcinoma. TNM staging of the tumors showed that seven were Stage IA, one Stage IB, one Stage IIA, and one Stage IIIA. Briefly, tunnel esophagogastrostomy is performed as follows: After performing a proximal gastrectomy, a rectangular seromuscular flap (3.0 cm × 3.5 cm) is created. The posterior esophageal wall is sutured to the gastric wall at the orad end of the seromuscular flap 5 cm from the stump with three to four stitches. Next, the stump of the esophagus is opened, the posterior esophageal wall is sutured to the gastric mucosa and submucosa, and the anterior esophageal wall is sutured to the full layer of the stomach. Finally, the caudad end of the seromuscular flap is closed. Data on surgical safety, postoperative morbidity, and postoperative reflux esophagitis were analyzed. All enrolled patients completed endoscopic follow-up 1 year and 2 years after surgery.Results:All procedures were completed. They comprised four cases of laparoscopic assisted surgery, four of DaVinci robotic surgery, and two of open surgery. The mean operation time was 212.7±33.2 mins, mean anastomosis time (51.6±5.3) minutes, mean tunnel preparation time (20.0±3.5) minutes, and mean operative blood loss (90.0±51.6) mL. The time to first postoperative passage of flatus was (64.8±11.5) hours. The mean hospital stay after surgery was (9.2±1.7) days. There were no postoperative complications above Clavien-Dindo Grade II. The mean preoperative Reflux Disease Questionnaire score was (3.3± 0.4) before the surgery, (3.8±1.0) 1 month postoperatively, and (3.3±0.4) 12 months postoperatively. All patients underwent endoscopic follow-up; no anastomotic stenoses were found. However, one patient had Grade A reflux esophagitis 1 year after surgery and another Grade B reflux esophagitis 2 years after surgery.Conclusion:Esophagogastrostomy by the tunnel technique is a safe and feasible means of performing proximal gastrectomy.

Cushing's disease is a severe endocrine disorder caused by excess secretion of adrenocorticotropic hormone from pituitary.Due to its subtle early clinical manifestations,the diagnosis of Cushing's disease is often delayed,and differentiating it from ectopic adrenocorticotropic hormone syndrome is a challenge.Combined functional tests can improve the diagnostic accuracy of Cushing's disease.The primary treatment for Cushing's disease is surgical removal of pituitary adenoma.However,about 1/3 of patients do not respond or experience recurrence after surgery,necessitating second-line treatments including medication.The development of novel drugs has enhanced the role of pharmacotherapy in the management of Cushing's disease.The review discusses the classic and emerging diagnostic methods and treatment strategies of Cushing's disease to deepen clinicians'understanding of its current treatment status and prospects.

Objective:To analyze the efficacy and feasibility of performing a new surgical procedure, tunnel esophagogastrostomy, to perform proximal gastrectomy.Methods:The study cohort comprised 10 consecutive patients who had undergone esophagogastrostomy by the tunnel technique in Jiangsu Cancer Hospital between October 2019 and July 2022. All patients were male. Their average age was (64.2±8.1) years and body mass index (25.5±3.2) kg/m2. Nine had upper gastric body adenocarcinoma, the remaining one having signet ring cell carcinoma. TNM staging of the tumors showed that seven were Stage IA, one Stage IB, one Stage IIA, and one Stage IIIA. Briefly, tunnel esophagogastrostomy is performed as follows: After performing a proximal gastrectomy, a rectangular seromuscular flap (3.0 cm × 3.5 cm) is created. The posterior esophageal wall is sutured to the gastric wall at the orad end of the seromuscular flap 5 cm from the stump with three to four stitches. Next, the stump of the esophagus is opened, the posterior esophageal wall is sutured to the gastric mucosa and submucosa, and the anterior esophageal wall is sutured to the full layer of the stomach. Finally, the caudad end of the seromuscular flap is closed. Data on surgical safety, postoperative morbidity, and postoperative reflux esophagitis were analyzed. All enrolled patients completed endoscopic follow-up 1 year and 2 years after surgery.Results:All procedures were completed. They comprised four cases of laparoscopic assisted surgery, four of DaVinci robotic surgery, and two of open surgery. The mean operation time was 212.7±33.2 mins, mean anastomosis time (51.6±5.3) minutes, mean tunnel preparation time (20.0±3.5) minutes, and mean operative blood loss (90.0±51.6) mL. The time to first postoperative passage of flatus was (64.8±11.5) hours. The mean hospital stay after surgery was (9.2±1.7) days. There were no postoperative complications above Clavien-Dindo Grade II. The mean preoperative Reflux Disease Questionnaire score was (3.3± 0.4) before the surgery, (3.8±1.0) 1 month postoperatively, and (3.3±0.4) 12 months postoperatively. All patients underwent endoscopic follow-up; no anastomotic stenoses were found. However, one patient had Grade A reflux esophagitis 1 year after surgery and another Grade B reflux esophagitis 2 years after surgery.Conclusion:Esophagogastrostomy by the tunnel technique is a safe and feasible means of performing proximal gastrectomy.

The detection principle of microfluidic microfluidic technology was introduced.The current research status of microfluidic platform-based SARS-CoV-2 nucleic acid detection technologies were reviewed such as reverse transcription quantitative real-time polymerase chain reaction(RT-qPCR),digital PCR,isothermal amplification and clustered regularly interspaced palindromic repeats/CRISPR-associated protein.The deficiencies of microfluidic platform-based SARS-CoV-2 nucleic acid detection were analyzed.It's pointed out microfluidic platform-based SARS-CoV-2 nucleic acid detection had to be optimized and validated clinically in specialty,sensitivity,detection limit,reproducibility,informatization,quality control and reagent cost.[Chinese Medical Equipment Journal,2024,45(1):101-107]

Objective:To explore the risk factors affecting the survival and efficacy of patients with acute myeloid leukemia with myelodysplasia-related changes(AML-MRC)transformed from myelodysplastic syndrome(MDS).Methods:The clinical data of 60 patients with AML-MRC transformed from MDS who hospitalized in The Third Affiliated Hospital of Soochow University from January 2010 to December 2021 were retrospectively analyzed.The demographic data and laboratory parameters,cytogenetic karyotypes,target genes of AML detected by next generation sequence,risk stratification,treatment regimen,therapeutic efficacy and survival outcome were documented.Rank sum test and Chi-square test or Fisher exact test were used to compare the survival and efficacy.The effects of clinical parameters,risk stratification and treatment regimens on the survival and efficacy of the AML-MRC patients were analyzed by univariate and multivariate analysis.Results:The median overall survival(OS)of the AML-MRC patients was 4.5 months,the 1-year OS rate was 28.3％,and the complete remission(CR)rate after treatment was 33.3％.The univariate analysis showed that age≥60 years,leukocytosis,severe thrombocytopenia,poor-risk group and only accepted hypomethylating agents(HMAs)or supportive therapy were the risk factors affecting OS.COX multivariate analysis showed that thrombocytopenia(HR=4.46),HMAs therapy(compared to transplantation,HR=10.47),supportive therapy(compared to transplantation,HR=25.80)and poor-risk group(compared to medium-risk group,HR=13.86)were independent hazard factors for median OS of patients with AML-MRC.The univariate analysis showed that the risk factors affecting 1-year OS in patients with AML-MRC were age ≥ 60 years,thrombocytopenia,time of transformation from MDS to AML(TTA)≥3 months,fibrinogen-albumin ratio index(FARI)≥ 0.07,CONUT score≥5,poor-risk group and supportive therapy.Binary logistic regression analysis showed that the independent risk factors for 1-year OS in AML-MRC patients were age ≥ 60 years(HR=11.23),thrombocytopenia(HR=8.71),FARI ≥ 0.07(HR=5.19)and poor-risk group(HR=14.00).The risk factors affecting CR of AML-MRC patients in univariate analysis were age ≥ 60 years,thrombocytopenia,FARI ≥ 0.1,CONUT score ≥ 5,poor-risk group and supportive therapy,while binary logistic regression analysis showed that age ≥ 60 years(HR=7.35),CONUT score ≥ 5(HR=9.60),thrombocytopenia(HR=12.05)and poor-risk group(HR=32.5)were independent risk factors affecting CR of the patients.Conclusion:The OS of AML-MRC patients is poor,old age(≥ 60 years old),supportive therapy,HMA therapy,poor-risk,thrombocytopenia,FARI ≥ 0.07 and CONUT score ≥ 5 may be associated with poor prognosis.

Objective:To investigate the prognostic significance of DTA(DNMT3A,TET2,ASXL1)gene mutations in patients with non-M3 acute myeloid leukemia(AML).Methods:The clinical data of 180 newly diagnosed AML patients hospitalized in the First People's Hospital of Changzhou from January 2018 to April 2022 were retrospectively analyzed.Next-generation sequencing technology was used to detect 150 gene mutations in the patients,and log-rank tests and Cox regression models were used to analyze the prognostic factors.Results:DTA gene mutations were detected in 83(46.1％)of 180 AML patients.Compared to patients without DTA mutations,patients with DTA mutations were significantly older(P＜0.001).The median overall survival(OS)time and disease-free survival(DFS)time in the DTA mutation group were significantly shorter than those in the group without DTA mutation(both P＜0.05).Multivariate analysis showed that age ≥ 60 years(P＜0.001),with DTA mutation(P=0.018),and intermediate-risk(relative to favorable-risk)(P=0.005)were independent risk factors for OS in AML patients.Conclusion:AML patients with DTA mutations are relatively older,with shorter median OS time and DFS time,and poor prognosis.

OBJECTIVE To investigate the current situation of the review and comment on inpatient medical orders in Beijing municipal hospitals， and to put forward countermeasures and suggestions for further improving related work. METHODS A questionnaire survey was conducted to investigate the current situation of the review and comment on inpatient medical orders in 22 Beijing municipal hospitals. The statistical analysis was conducted for the survey results. RESULTS A total of 22 questionnaires were distributed， with recovery effective rate of 100%. The 22 hospitals carried out inpatient medical order comment， but their proportion varied among hospitals （0.88%-98.54%）； medical order comment mainly focused on antibiotics， proton pump inhibitors， anesthetic drugs/class Ⅰ psychotropic drugs， auxiliary drugs and other categories； 205 pharmacists participated in the comment of inpatient medical orders， most of whom hold intermediate or higher professional titles （89.27%）； 21 hospitals conducted inpatient medical order comment and feedback the results to relevant departments/responsible persons， but the intervention situation was not the same. Eighteen hospitals had carried out the review of inpatient medical orders； reviewed drug category was roughly the same as the category involved in the medical order comment； review content involved the suitability of administration routes， dosage， etc. The review was conducted mainly through the cooperation of audit software and pharmacists. CONCLUSIONS The comment and review of inpatient medical orders in Beijing municipal hospitals carried out in an orderly manner， and preliminary results have been achieved； at the same time， it is necessary to further increase the ability of participating pharmacists， improve audit standards， optimize pre-audit （No.ZYLX201805） software， and promote rational drug use among hospitalized patients through doctor-pharmacist collaboration.

[Abstract] [Objective] To construct inducible immortalization gene vectors for transfection into primary cells, enabling the establishment of a conditionally immortalized cell line that support their sustained cultivation and proliferation in vitro. [Methods] Using gene homologous recombination technology, the coding sequences (CDS) of immortalization genes-including human telomerase reverse transcriptase (hTERT), simian virus 40 large T antigen (SV40LT), acute myeloid leukemia fusion genes NUP98-KDM5A (N/K) and CBFA2T3-GLIS2 (C/G), as well as the proto-oncogene KRAS were precisely inserted into the tetracycline (Tet)-inducible eukaryotic expression lentiviral vector pLV2-TRE3GS-EGFP-MCS-3×FLAG-hPGK-Tet-On-SV40-Neo and the transposon PB-TRE3G-3×FLAG-T2A-Puro-SV40-PA. Lentiviral packaging, cell transfection, mRNA expression analysis, Western blotting for protein detection, green fluorescent protein (GFP) visualization, and cell proliferation assays were conducted to evaluate transfection efficiency and assess the regulatory effects of Tet on gene expression in 293T and MEF cells. [Results] The Tet-inducible lentiviral vectors pLV2-Tet-SV40LT, pLV2-Tet-N/K, and pLV2-Tet-C/G, along with the transposon vectors PB-Tet-hTERT, PB-Tet-SV40LT, PB-Tet-N/K, PB-Tet-C/G, and PB-Tet-KRAS, were successfully constructed. In 293T cells, the expression levels of all target genes were upregulated after transfection. In MEF cells, the immortalizing functions of SV40LT and N/K were validated. By modulating Tet addition, cell proliferation levels were effectively regulated, leading to the successful establishment of conditionally immortalized pLV2-SV40LT-MEF and pLV2-N/K-MEF cell lines. [Conclusion] The construction of Tet-inducible immortalizing gene vectors provides a technical foundation for establishing conditionally immortalized primary cell lines, thereby facilitating research on the large-scale in vitro production and expansion of blood cells, such as erythrocytes and platelets.