OBJECTIVE To establish a preparation system for megakaryocytes(MKs)derived from human embryonic stem cells(hESCs)and MK microparticles(MKMPs),and to assess the pro-angio-genic efficiency of these microparticles.METHODS ①hESCs were induced to mesodermal progenitor cells via monolayer culture with the first-stage induction medium for 2 days before the cells were induced to hemogenic endothelial/hematopoietic progenitor cells by culturing with the second-stage induction medium for another 3 days.Then,the cells were dissociated into single cells,seeded into the third-stage induction medium,and cultured using the suspension method for 8 days to obtain MKs.The specific characters of differentiated cells were identified through morphological observation and flow cytometry before stage-specific marker proteins in different periods were analyzed[hESCs:TRA-1-60,sialyl glycolipid stage-specific embryonic antigen4(SSEA4)];mesodermal progenitor cells:brachyury;hemogenic endothelial/hematopoietic progenitor cells:CD34,CD43;MKs:CD41a,CD42b),and immu-nofluorescence staining[β1-tubulin,von Willebrand factor(VWF)],[friend leukemia integration 1(FLI1),CD42].② MKMP collection and verification:MKMPs were collected via differential centrifugation.The concentration and size of these MKMPs were determined by nanoparticle tracking analysis(NTA),and both the morphology and ultrastructure were examined by transmission electron microscopy(TEM).Besides,the MKMPs-specific proteins[CD41,tumor susceptibility gene 101(TSG101)and CD9]were detected by Western blotting analysis.③ Biological function of MKMPs:MKMPs were stained with CD41a-PE antibodies and co-cultured with human umbilical veinvascular endothelial cells(HUVECs)labeled by CD34-APC for 3 h.Live-cell immunofluorescence was employed to find out whether HUVECs could absorb MKMPs.To find out whether MKMPs could affect the role of HUVECs in angio-genesis and cell migration,platelet microvesicles(PMPs)were used as positive controls.The experi-mental groups were added with different concentrations of microparticles(1,5,10 and 20 mg·L-1)while the control group was given no microparticles(0 mg·L-1).The number of nodes that formed the lumen after 5 h of incubation in Matrigel was counted,and the size of healing of the scratch area was analyzed after 6 h.To elucidate the mechanism through which MKMPs impacted angiogenesis,ELISA was used out to quantitatively detect the concentration of proteins in microparticles.RESULTS ① A three-stage differentiation cultural system was established to develop hESCs into MKs.Flow cytometry revealed progressive loss of pluripotency markers SSEA4 and TRA-1-60,while the mesodermal progenitor marker brachyury peaked at d 2.Subsequently,hemogenic endothelial/hematopoietic progenitor markers CD34 and CD43 emerged at d 5,followed by megakaryocytic markers CD41a and CD42b at d 13.Immunofluorescent images further demonstrated that MKs expressed specific proteins CD42,β1-tubulin,von VWF and FLI1 at d 13.②Microparticles were collected via differential centrifuga-tion.Transmission electron microscopy revealed that their substructure exhibited a typical double-layered membrane.Nanoparticle tracking analysis indicated that the size was(164.3±14.0)nm.The result of WB demonstrated that the microparticles expressed specific markers,including TSG101,CD9 and CD41.③ MKMPs were absorbed after being co-cultured with HUVECs for 3 h and enhanced the ability of HUVECs to form tubes and migrate.Notably,the treatment of 5 mg·L-1 MKMPs was more effective than 5 mg·L-1 PMPs treatment.The results of ELISA showed that the content of VEGF from MKMPs was higher than from PMPs,which may be the key factor in regulating endothelial biological function.CONCLUSION MKs derived from hESCs can generate functional microparticles which can promote angiogenesis.

OBJECTIVE To establish a preparation system for megakaryocytes(MKs)derived from human embryonic stem cells(hESCs)and MK microparticles(MKMPs),and to assess the pro-angio-genic efficiency of these microparticles.METHODS ①hESCs were induced to mesodermal progenitor cells via monolayer culture with the first-stage induction medium for 2 days before the cells were induced to hemogenic endothelial/hematopoietic progenitor cells by culturing with the second-stage induction medium for another 3 days.Then,the cells were dissociated into single cells,seeded into the third-stage induction medium,and cultured using the suspension method for 8 days to obtain MKs.The specific characters of differentiated cells were identified through morphological observation and flow cytometry before stage-specific marker proteins in different periods were analyzed[hESCs:TRA-1-60,sialyl glycolipid stage-specific embryonic antigen4(SSEA4)];mesodermal progenitor cells:brachyury;hemogenic endothelial/hematopoietic progenitor cells:CD34,CD43;MKs:CD41a,CD42b),and immu-nofluorescence staining[β1-tubulin,von Willebrand factor(VWF)],[friend leukemia integration 1(FLI1),CD42].② MKMP collection and verification:MKMPs were collected via differential centrifugation.The concentration and size of these MKMPs were determined by nanoparticle tracking analysis(NTA),and both the morphology and ultrastructure were examined by transmission electron microscopy(TEM).Besides,the MKMPs-specific proteins[CD41,tumor susceptibility gene 101(TSG101)and CD9]were detected by Western blotting analysis.③ Biological function of MKMPs:MKMPs were stained with CD41a-PE antibodies and co-cultured with human umbilical veinvascular endothelial cells(HUVECs)labeled by CD34-APC for 3 h.Live-cell immunofluorescence was employed to find out whether HUVECs could absorb MKMPs.To find out whether MKMPs could affect the role of HUVECs in angio-genesis and cell migration,platelet microvesicles(PMPs)were used as positive controls.The experi-mental groups were added with different concentrations of microparticles(1,5,10 and 20 mg·L-1)while the control group was given no microparticles(0 mg·L-1).The number of nodes that formed the lumen after 5 h of incubation in Matrigel was counted,and the size of healing of the scratch area was analyzed after 6 h.To elucidate the mechanism through which MKMPs impacted angiogenesis,ELISA was used out to quantitatively detect the concentration of proteins in microparticles.RESULTS ① A three-stage differentiation cultural system was established to develop hESCs into MKs.Flow cytometry revealed progressive loss of pluripotency markers SSEA4 and TRA-1-60,while the mesodermal progenitor marker brachyury peaked at d 2.Subsequently,hemogenic endothelial/hematopoietic progenitor markers CD34 and CD43 emerged at d 5,followed by megakaryocytic markers CD41a and CD42b at d 13.Immunofluorescent images further demonstrated that MKs expressed specific proteins CD42,β1-tubulin,von VWF and FLI1 at d 13.②Microparticles were collected via differential centrifuga-tion.Transmission electron microscopy revealed that their substructure exhibited a typical double-layered membrane.Nanoparticle tracking analysis indicated that the size was(164.3±14.0)nm.The result of WB demonstrated that the microparticles expressed specific markers,including TSG101,CD9 and CD41.③ MKMPs were absorbed after being co-cultured with HUVECs for 3 h and enhanced the ability of HUVECs to form tubes and migrate.Notably,the treatment of 5 mg·L-1 MKMPs was more effective than 5 mg·L-1 PMPs treatment.The results of ELISA showed that the content of VEGF from MKMPs was higher than from PMPs,which may be the key factor in regulating endothelial biological function.CONCLUSION MKs derived from hESCs can generate functional microparticles which can promote angiogenesis.

OBJECTIVE To assess the long-term cost-effectiveness of five glucagon-like peptide-1 receptor agonists （GLP- 1RAs） in the treatment of poorly controlled type 2 diabetes mellitus （T2DM） treated with metformin. METHODS Baseline data from patients in previously published meta-analysis and included randomized controlled trials （RCTs） were extracted to predict survival， long-term efficacy， and costs for each group using the United Kingdom prospective diabetes study outcome model 2.1. The cost-effectiveness of 5 GLP-1RAs （liraglutide， lixisenatide， exenatide， dulaglutide， and semaglutide） was analyzed by cost- utility analysis. Sensitivity analysis and scenario analysis were also performed to verify the uncertainty of basic analysis results. RESULTS A total of 21 RCTs with 6 796 patients were included. Survival analysis curves showed the superiority of semaglutide in reducing the risk of death from cardiovascular disease and dulaglutide in reducing the risk of all-cause mortality over other GLP- 1RAs. The cost-utility analysis showed that the five drugs were economically superior to inferior in the order of lixisenatide， semaglutide， exenatide， dulaglutide， and liraglutide； one-way and probabilistic sensitivity analyses indicated that the results were robust. The scenario analysis results indicated that the price of semaglutide should decrease by at least 54.64% to 369.21 yuan， which is cost-effectiveness compared to lixisenatide. CONCLUSIONS For T2DM patients in China with poor glycemic control after treatment with metformin， lixisenatide and semaglutide may be considered as the preferred regimen.

Objective:To investigate the clinical efficacy of 3D printed metal augment or tibial prosthesis for reconstruction of large bone defects in total knee arthroplasty (TKA) and knee revision surgery.Methods:A total of 7 patients (7 knees) with TKA or knee revision who were admitted to the Department of Orthopaedics of the Second Affiliated Hospital of Zhejiang University School of Medicine with large bone defects from July 2018 to December 2023 were retrospectively analyzed, including 4 patients with TKA and 3 patients with knee revision. There were 3 males and 4 females, aged 58.7±7.6 years (range, 54-68 years), 3 patients with left knee and 4 patients with right knee. All the patients had bone defects in the knee joint (AORI type III), 2 cases had bone defects only in the femur, 4 cases had bone defects only in the tibia, and 1 case had bone defects in both the tibia and femur, which were treated with personalized reconstruction using 3D printing. Hip-knee-ankle angles, American Knee Society score (KSS) before and after surgery were compared, and postoperative complications were observed.Results:All patients successfully completed the operation, and the operation time was 189.3±35.5 min (range, 125-240 min). Complex TKA was performed in 4 cases with surgical times of 175, 195, 210, and 240 min, and revision surgery was performed in 3 cases with surgical times of 125, 180, and 200 min, respectively. Intraoperative blood loss was 114±24.4 ml (range, 100-150 ml). Five cases used 3D printed metal augment, and two used 3D printed one-piece tibial components. All patients were followed up for 2, 2, 5, 6, 7, 20, 57 months, respectively. The KSS of the five patients at 3 months postoperatively were 56, 61, 66, 56, and 56 points, respectively, greater than the preoperative scores of 35, 44, 36, 27, and 41 points. The KSS functional scores of the five patients at 3 months postoperatively were 45, 45, 45, 30, and 45 points, respectively, which were greater than the preoperative scores of 30, 30, 15, 20, and 20 points. The hip-knee-ankle angle was 181.8°±3.4° (range, 177.9° to 188.0°) at the final follow-up and 175.8°±12.4° (range, 153.3° to 192.1°) before surgery, with no significant difference ( t=-1.230, P=0.242). At the final follow-up, the 3D printed component was well integrated with the bone surface, the prosthesis was securely positioned, and the force lines of the lower limbs were normal. There were no postoperative complications such as poor wound healing, infection, fat liquefaction, nerve injury, deep vein thrombosis of lower limbs, knee joint stiffness, periprosthesis infection and loosening. Conclusion:Using 3D printed metal augment or tibial prosthesis to reconstruct the huge bone defect in TKA and revision has a satisfactory early clinical effect, satisfactory joint function and good surgical safety.

Severe acute pancreatitis(SAP)is a critical condition in general surgery settings,characterized by high mortality and poor prognosis.On February 28,2024,the Department of Hepatobiliary Surgery at the First Affiliated Hospital of Harbin Medical University admitted a 36-year-old male patient.The patient presented with"upper abdominal pain accompanied by fever for three months and jaundice of the skin and sclera for one week."Physical examination revealed 11 puncture tubes,and a palpable mass measuring 3 cm × 5 cm in the upper abdomen.Enhanced CT and magnetic resonance cholangiopancreatography indicated acute pancreatitis.The patient was diagnosed with"SAP,infectious pancreatic necrosis,and biliary stenosis."He had severe abdominal infection and complex postoperative complications,making treatment challenging.Consequently,a multidisciplinary team(MDT)consultation was initiated.After three rounds of MDT consulfation and freating,the patient ultimately recovered successfully and was discharged.This article reviews the MDT treatment process for this patient and summarizes the characteristics of this condition based on relevant literature to provide insights and experience for clinical practice.

OBJECTIVE To evaluate the effectiveness and safety of trastuzumab biosimilars and original drugs in the treatment of human epidermal growth factor receptor -2（HER-2）positive breast cancer systematically ，and to provide evidence -based basis for the selection of clinical treatment . METHODS Retrieved from PubMed ，Embase，Web of Science ，Cochrane Library ，CNKI， CBM，VIP，Wanfang database ，ClinicalTrials and Clinical Trial Center of China ，randomized controlled trials （RCTs）about trastuzumab biosimilars （trial group ）versus original drugs （control group ）in the treatment of HER -2 positive breast cancer were collected. After data extraction and quality evaluation ，RevMan 5.4 statistical software was used for meta -analysis. RESULTS Totally 11 RCTs were enrolled ，involving 5 812 patients. Results of meta -analysis showed that there was no significant difference in objective response rate （based on whether intention -to-treat population nor per -protocol population ），pathologic complete response ， proportion of progression free survival population ，proportion of overall survival population ，anti-drug antibody positive rate ， neutralizing antibody positive rate ，and incidence of adverse drug reactions between 2 groups（P＞0.05）. CONCLUSIONS The effectiveness and safety of trastuzumab biosimilars are equivalent to those of trastuzumab original drugs .

Objective:To explore the clinical features, treatment and prognosis of focal nodular hyperplasia (FNH) of the liver.Methods:A retrospective analysis of the clinical data of 36 patients with FNH who had undergone surgery and were pathologically confirmed from May 2013 to Aug 2021 was made at the General Surgery Department of the First Affiliated Hospital of Nanchang University.Results:There were 13 males and 23 females, with an average age of (35.9±15.3) years. 50% patients were asymptotic,21 cases (58.3%) were diagnosed as FNH by imaging before operation. All patients underwent surgical resection. There was no postoperative mortality.Pathology confirmed preoperative tentative diagnosis of FNH in all cases. The average follow-up time was 21.1 months, all patients were alive, and 1 patient had a relapse.Conclusions:FNH is a benign liver tumor like lesion. Surgery is suggested for cases suspected of tumor and the prognosis is good.

A 9-year-old boy presented with pruritic erythema and erosions of the scalp for 1 month, which spread all over the body for 1 week. Histopathological examination of the skin lesions showed blisters forming under the granular layer of the epidermis, a large number of acantholytic cells in the blisters, and perivascular infiltration of lymphocytes and eosinophils in the superficial dermis. Direct immunofluorescence study showed reticular deposition of IgG and complement C3 between epidermal cells, and negative staining for IgM and IgA. Enzyme-linked immunosorbent assay showed the presence of serum anti-Dsg1 antibody (157.00 U/ml) . The patient was diagnosed with pemphigus foliaceus. After admission, the patient showed poor response to the treatment with prednisone at a dose of 40 mg/d. After the treatment with low-dose rituximab (100 mg per week, 4 weeks) combined with prednisone (20 mg/d) , her condition was well controlled. The dosage of prednisone gradually decreased to 7.5 mg/d for maintenance treatment, and no recurrence was observed during 24-month follow-up.

Objective To summarize the clinical characteristics of autoimmune pancreatitis (AIP) and discuss the role and significance of surgical intervention in the treatment of AIP.Methods The clinical data of 11 patients with preoperative confirmed AIP and 8 patients with preoperative suspected AIP receiving surgical intervention at the First Affiliated Hospital of Harbin Medical University from 2011 to 2017 were retrospectively analyzed.Results Among 11 preoperatively confirmed AIP patients,9 (47.4％) underwent endoscopic retrograde cholangiopancreatography (ERCP) with nasal bile duct implantation or biliary stent drainage,and 2 (10.5％)underwent choledochojejunostomy.Among 8 putative AIP patients in which a cancer was suspected 4 (21.1％) underwent choledochojejunostomy,3 (15.8％) did pancreaticoduodenectomy,and 1 (5.3％) had distal pancreatectomy plus splenectomy.Postoperative pathology confirmed pancreatic cancer in 3 cases,IPMN in 1 case.Conclusions While autoimmune pancreatitis is IgG4 related disease,surgical intervention is indicated when there is poor response to drug treatment or when a cancer is to be ruled out.

With the introduction and development of the concept of precision surgery,3D printing individualized guide template has been gradually applied in the field of spinal surgery because of its advantages of increasing surgical precision,reducing the difficulty and risk of surgery,reducing the operation time,and reducing the intraoperative transmission dose,etc..Individualized guide plates designed and manufactured according to the patient~ surgical site,self-conditions and individual differences before surgery can accurately restore the preoperative design during surgery to meet the needs of modernized spine surgery for individualization and precision.Individualized guide template has great guiding significance for complex spinal surgery.This article reviews the current clinical application of 3D printing individualized guide template in spine surgery,summarizes its produce principles and procedures,advantages and limits,and make prospect for it's future applications.