Heterozygous Prothrombin Mutation-Associated Thrombophilia.

BACKGROUND: Venous thromboembolism (VTE) is predisposed by thrombotic mutations in patients with hereditary thrombophilia. Although prothrombin deficiencies caused by homozygous or compound heterozygous mutations are associated with bleeding diathesis, rare cases have shown a correlation between heterozygous prothrombin mutations and thrombosis. MATERIALS AND METHODS: We surveyed genetic variants involved in thrombosis and hemostasis in 347 patients with unprovoked VTE or having a positive family history of thrombosis. For patients identified with heterozygous prothrombin mutations, we conducted family investigations and performed a thrombin generation test (TGT) to elucidate the thrombotic risk. Novel mutants were expressed and subjected to functional assays to clarify the underlying thrombotic mechanisms. RESULTS: Heterozygous prothrombin mutations were identified in 3.5% of patients (12/347), including three novel mutations Phe382Ser, Phe382Leu, and Asp597Tyr found in one patient each, as well as previously reported Arg541Trp mutation in four patients and Arg596Gln mutation in five patients. A total of 42 mutation carriers were identified within the 12 pedigrees, among whom 64.3% (27/42) had experienced thrombotic events. TGT results demonstrated hypercoagulability for carriers of the five mutations, with Arg596Gln showing the highest thrombin generation potential followed by Arg541Trp. The Phe382-associated mutations severely impaired thrombomodulin-binding ability of thrombin, resulting in obviously reduced protein C (PC) activation. The Asp597Tyr mutation exhibited a mild reduction in both inactivation by antithrombin and PC activation reactions. CONCLUSION: The presence of heterozygous prothrombin mutations represents a potential genetic predisposition for VTE. All thrombosis-associated mutations potentiate coagulation activity by either conferring antithrombin resistance and/or impairing PC pathway activity.

Intratumoral CXCR4hi neutrophils display ferroptotic and immunosuppressive signatures in hepatoblastoma.

Pediatric hepatoblastoma (HB) is the most common primary liver malignancy in infants and children. With great diversity and plasticity, tumor-infiltrating neutrophils were one of the most determining factors for poor prognosis in many malignant tumors. In this study, through bulk RNA sequencing for sorted blood and tumor-infiltrated neutrophils and comparison of neutrophils in tumor and para-tumor tissue by single-cell sequencing, we found that intratumoral neutrophils were composed of heterogenous functional populations at different development stages. Our study showed that terminally differentiated neutrophils with active ferroptosis prevailed in tumor tissue, whereas, in para-tumor, pre-fate naïve neutrophils were dominant and ferroptotic neutrophils dispersed in a broad spectrum of cell maturation. Gene profiling and in vitro T-cell coculture experiment confirmed that one of main functional intratumoral neutrophils was mainly immunosuppressive, which relied on the activation of ferroptosis. Combining the bulk RNA-seq, scRNA-seq data, and immunochemistry staining of tumor samples, CXCL12/CXCR4 chemotaxis pathway was suggested to mediate the migration of neutrophils in tumors as CXCR4 highly expressed by intratumoral neutrophils and its ligand CXCL12 expressed much higher level in tumor than that in para-tumor. Moreover, our study pinpointed that infiltrated CXCR4hi neutrophils, regardless of their differential distribution of cell maturation status in HB tumor and para-tumor regions, were the genuine perpetrators for immune suppression. Our data characterized the ferroptosis-dependent immunosuppression energized by intratumoral CXCR4 expression neutrophils and suggest a potential cell target for cancer immunotherapies.

Elevated exosome-transferrable lncRNA EPB41L4A-AS1 in CD56bright NK cells is responsible for the impaired NK function in neuroblastoma patients by suppressing cell glycolysis.

NK cells are one of key immune components in neuroblastoma (NB) surveillance and eradication. Glucose metabolism as a major source of fuel for NK activation is exquisitely regulated. Our data revealed a diminished NK activation and a disproportionally augmented CD56bright subset in NB. Further study showed that NK cells in NB presented with an arrested glycolysis accompanied by an elevated expression of the long noncoding RNA (lncRNA) EPB41L4A-AS1, a known crucial participant in glycolysis regulation, in the CD56bright NK subset. The inhibitory function of lncRNA EPB41L4A-AS1 was recapitulated. Interestingly, our study demonstrated that exosomal lncRNA EPB41L4A-AS1 was transferrable from CD56bright NK to CD56dim NK and was able to quench the glycolysis of target NK. Our data demonstrated that an arrested glycolysis in patient NK cells was associated with an elevated lncRNA in CD56bright NK subset and a cross-talk between heterogeneous NK subsets was achieved by transferring metabolic inhibitory lncRNA through exosomes.

Esophageal Carcinosarcoma: Analysis of Clinical Features and Prognosis of 24 Cases and a Literature Review.

OBJECTIVE: Esophageal carcinosarcoma (ECS) is a rare malignant tumor that accounts for only 0.5%-2.8% of all esophageal malignancies. As most current studies are case reports, the relationship between clinical features and prognosis remains controversial. METHODS: We investigated the clinical features and prognosis of 24 patients with ECS in a single center from 2006 to 2018. There were 18 male and 6 female patients aged 52-82 years with a median age of 62.5 years. In addition, we included 9 studies on ECS from PubMed and a literature review. RESULTS: The median follow-up time of the 24 patients was 70.5 (range, 10-156)months. The 3-year and 5-year survival rates were 83.3% and 70.8%, respectively. Among the 24 patients, none of the 10 (41.7%) stage T1 cancer patients had lymph node metastasis; however, lymph node metastasis was noted in 8 (57.1%) stage T2-4 cancer patients. The literature review revealed that 211 patients had a 5-year survival rate of 11.8%-68.2%, and 54.5%-95.8% study participants had early stage ECS. Although the information provided in the literature review is limited, it appears to be a characteristic of the early stage of the disease and predicts better prognosis when ECS is diagnosed, which is similar to the result of the current study. CONCLUSION: Our results indicate that ECS has a favorable prognosis, even among patients with early stage ECS who undergo radical esophagectomy with lymph node dissection. Because of the low incidence of ECS, further studies with more cases need to investigate this rare malignancy.

Identification of two novel mutations in three children with congenital factor VII deficiency.

Congenital factor VII deficiency (FVIID) is a rare F7 gene mutation causing bleeding disorder inherited in an autosomal recessive manner. In this study, we aimed to identify genetic defects and analyze their relationships with phenotype in three Chinese FVIID patients. The diagnosis of FVIID was made based on FVII coagulant activity (FVII:C) levels assessed through prothrombin time assay. Direct sequencing and protein modeling were performed to detect genetic mutations and the resulting protein expression. Patient 1, a 2-year-old girl, presented with mild bleeding and was found to have a FVII:C of 0.2% and a compound heterozygous F7 Cys389Gly/Cys115Arg mutation. Patient 2, a 7-year-old boy, consulted for moderate bleeding and was found to have a FVII:C of 0.8% and a compound heterozygous F7 Thr241Asn/Pro324Leu mutation. Patient 3, a 5-year-old boy who developed a mild bleeding after trauma was found to have a FVII:C of 1.8% and a compound heterozygous F7 Thr241Asn/ IVS5-2A>G mutation. We hereby report three congenital FVIID patients with FVII:C less than 2% and their respective F7 mutations, two of which (F7 Cys115Arg, Pro324Leu) are novel. The molecular model analysis of the two novel mutations F7 Cys115Arg and Pro324Leu respectively indicated impairment of the proper folding of epidermal growth factor 1 domain situated on F7 gene and impairment of the procoagulant function of FVII both leading to the congenital deficiency of FVII.

Identification of one novel pathogenic ITGB3 mutation and two known mutations in two Chinese pedigrees with hereditary Glanzmann thrombasthenia.

Glanzmann thrombasthenia (GT) is an inherited disorder of platelet aggregation resulting from quantitative and/or qualitative abnormalities of the glycoprotein IIb/IIIa complex. We analyzed the expression of GPIIb/IIIa and the gene sequencing in two pedigrees with GT, so as to determine the type and the relationship between genotype and clinical phenotype. Platelet aggregation tests and flow cytometric studies were performed, along with gene sequencing. Both probands were classified as grade III of bleeding. Platelet aggregation was absent or defective upon stimulation with physiological stimuli like AA and ADP, but platelets agglutinated normally in response to ristocetin. MFI values were considerably reduced. Gene sequencing showed ITGB3 mutations p.Cys549Ser/p.Leu705CysfsTer4 in proband 1 and p.Cys549Ser/p.Gln254Lys in proband 2 and her sister. This study reports one novel ITGB3 mutant gene, p.Gln254Lys, of which we will explore the potential pathogenicity.