Expression of angiostatin cDNA in human gallbladder carcinoma cell line GBC-SD and its effect on endothelial proliferation and growth

AIM： To explore the influence of angiostatin up-regulation on the biologic behavior of gallbladder carcinoma cells in vitro and in vitro, and the potential value of angiostatin gene therapy for gallbladder carcinoma. METHODS： A eukaryotic expression vector of pcDNA3.1（＋） containing murine angiostatin was constructed and identified by restriction endonuclease digestion and sequencing. The recombinant vector pcDNA3.1-angiostatin was transfected into human gallbladder carcinoma cell line GBC-SD with Upofectamine 2000, and paralleled with the vector and mock control. The resistant clone was screened by G418 filtration. Angiostatin transcription and protein expression were examined by RT-PCR, immunofluorescence and Western-blot. The supernatant was collected to treat endothelial cells. Cell proliferation and growth in vitro were observed under microscope. RESULTS： Murine angiostatin cDNA was successfully cloned into the eukaryotic expression vector pcDNA3.1 （＋）. AFter 14 d of transfection and selection with G418, macroscopic resistant cell cloning was formed in the experimental group transfected with pcDNA 3.1（＋）-angiostatin and vector control. But untreated cells died in the mock control. Angiostatin was detected by RT-PCR and protein expression was detected in the experimental group by immunofluorescence and Western-blot. Cell proliferation and growth in v/tro in the three groups were observed respectively under microscope. No significant difference was observed in the growth speed of GBC- SD cells between groups that were transfected with and without angiostatin. After treatment with supernatant, significant differences were observed in endothelial cell （ECV-304） growth in vitro. The cell proliferation and growth were inhibited. CONCLUSION： Angiostatin does not directly inhibit human gallbladder carcinoma cell proliferation and growth in vitro, but the secretion of angiostatin inhabits endothelial cell proliferation and growth.

Angiostatin inhibits pancreatic cancer cell proliferation and growth in nude mice

AIM: To observe the biologic behavior of pancreatic cancer cells in vitro and in vivo, and to explore the potential value of angiostatin gene therapy for pancreatic cancer.METHODS: The recombinant vector pcDNA3.1(+)-angiostatin was transfected into human pancreatic cancer cells PC-3 with Lipofectamine 2000, and paralleled with the vector and mock control. Angiostatin transcription and protein expression were determined by immunofluorescence and Western blot. The stable cell line was selected by G418. The supernatant was collected to treat endothelial cells. Cell proliferation and growth in vitro were observed under microscope. Cell growth curves were plotted.The troms-fected or untroms-fected cells overexpressing angiostatin vector were implanted subcutaneously into nude mice. The size of tumors was measured, and microvessel density count (MVD) in tumor tissues was assessed by immunohistochemistry with primary anti-CD34antibody.RESULTS: After transfected into PC-3 with Lipofectamine 2000 and selected by G418, macroscopic resistant cell clones were formed in the experimental group transfected with pcDNA 3.1(+)-angiostatin and vector control. But untreated cells died in the mock control. Angiostatin protein expression was detected in the experimental group by immunofluorescence and Western-blot. Cell proliferation and growth in vitro in the three groups were observed respectively under microscope. After treatment with supernatant, significant differences were observed in endothelial cell (ECV-304) growth in vitro. The cell proliferation and growth were inhibited. In nude mice model, markedly inhibited tumorigenesis and slowed tumor expansion were observed in the experimental group as compared to controls, which was parallel to the decreased microvessel density in and around tumor tissue.CONCLUSION: Angiostatin does not directly inhibit human pancreatic cancer cell proliferation and growth in vitro,but it inhibits endothelial cell growthin vitro. It exerts the anti-tumor functions through antiangiogenesis in a paracrine way in vivo.

Combining Serum Procalcitonin Level,Thromboelastography,and Platelet Count to Predict Short-term Development of Septic Shock in Intensive Care Unit

Objective Despite the recent advances in diagnosis and treatment,sepsis continues to lead to high morbidity and mortality.Early diagnosis and prompt treatment are essential to save lives.However,most biomarkers can only help to diagnose sepsis,but cannot predict the development of septic shock in high-risk patients.The present study determined whether the combined measurement of procalcitonin(PCT),thromboelastography(TEG)and platelet(PLT)count can predict the development of septic shock.Methods A retrospective study was conducted on 175 septic patients who were admitted to the intensive care unit between January 2017 and February 2021.These patients were divided into two groups:73 patients who developed septic shock were assigned to the septic shock group,while the remaining 102 patients were assigned to the sepsis group.Then,the demographic,clinical and laboratory data were recorded,and the predictive values of PCT,TEG and PLT count for the development of septic shock were analyzed.Results Compared to the sepsis group,the septic shock group had statistically lower PLT count and TEG measurements in the R value,K value,αangle,maximum amplitude,and coagulation index,but had longer prothrombin time(DT),longer activated partial thromboplastin time(APTT),and higher PCT levels.Furthermore,the Sequential Organ Failure Assessment(SOFA)score was higher in the septic shock group.The multivariate logistic regression analysis revealed that PCT,TEG and PLT count were associated with the development of septic shock.The area under the curve analysis revealed that the combined measurement of PCT,TEG and PLT count can be used to predict the development of septic shock with higher accuracy,when compared to individual measurements.Conclusion The combined measurement of PCT,TEG and PLT count is a novel approach to predict the development of septic shock in high-risk patients.

Anlotinib in combination with pembrolizumab for low-grade myofibroblastic sarcoma of the pancreas: A case report

BACKGROUND Low-grade myofibroblastic sarcoma(LGMS)is a rare spindle cell sarcoma espe-cially in the pancreas,with myofibroblastic differentiation.Hitherto,only a few cases have been reported.CASE SUMMARY Herein,we report a case involving the discovery of a pancreatic mass detected during a routine physical examination.Subsequent imaging and pathological tests of the patient led to the diagnosis of LGMS of the pancreas.Following surgical intervention,the patient experienced recurrence and metastasis.Conventional treatment is not effective for postoperative recurrent pancreatic LGMS with multiple metastases.After communicating with the patients and their families,informed consent was obtained for the treatment of anlotinib combined with pembrolizumab.Evaluation of imaging and clinical symptoms post-treatment revealed a relatively favorable response to the combination of anlotinib and pembrolizumab.CONCLUSION Based on the comprehensive literature review,our report aimed to provide evidence for a better understanding of the disease characteristics,diagnostic criteria,imaging findings,and identification of LGMS.And explore novel treatment strategies for this disease.