This study explores the effects of enhancing the definitive hematopoiesis(DH)signals during the differentiation of human embryonic stem cells(hESCs)into hematopoietic stem/progenitor cells on the generation efficiency and effector function of natural killer(NK)cells generated from hESCs(also known as hESC-NK cells).The hESC(H1)were transformed into embryoid bodies(Ebs)by centrifugation,and during the induction of K562,were used to analyze the efficiency of hematopoietic differentiation,the efficiency of NK cell generation from hESC,the in vitro effector functions,and the expression of effector function related surface receptors.Compared to the control group,the DH group had a significant increase in the number of arterial hematopoietic endothelial cells(CD34+DLL4+)and a significant decrease in primitive hematopoietic related cells(CD34-CD43+)on day 8 of hematopoietic differentiation(P<0.05).On day 28 of NK cell differentiation,the DH group demonstrated a significant increase in the number of NK cells(CD45+CD56+),while a slight increase in the expression of effector function-related molecules such as IFN-γ,Granzyme B,Perforin and CD107a without statistical significance.Furthermore,the activation receptors CD16a and CD69 were significantly increased,NKP46 was significantly decreased,the inhibitory receptor NKG2A was significantly increased,while CD96 was significantly decreased on hESC-NK cells of DH groups(P<0.05).Conclusively,enhancing the signals for definitive hematopoiesis during hESC differentiation into hematopoietic stem/progenitor cells significantly improves the yield of NK cells and the expression of CD16a without affecting their in vitro effector functions.Our study provides a new approach to improving the efficiency of hESC-NK cell or iPSC-NK cell generation.

Objective: To compare the incidence of radiation-related toxicities between conventional and hypofractionated intensity-modulated radiation therapy (IMRT) for limited-stage small cell lung cancer (SCLC), and to explore the risk factors of hypofractionated radiotherapy-induced toxicities. Methods: Data were retrospectively collected from consecutive limited-stage SCLC patients treated with definitive concurrent chemoradiotherapy in Cancer Hospital of Chinese Academy of Medical Sciences from March 2016 to April 2022. The enrolled patients were divided into two groups according to radiation fractionated regimens. Common Terminology Criteria for Adverse Events (CTCAE, version 5.0) was used to evaluate the grade of radiation esophagus injuries and lung injuries. Logistic regression analyses were used to identify factors associated with radiation-related toxicities in the hypofractionated radiotherapy group. Results: Among 211 enrolled patients, 108 cases underwent conventional IMRT and 103 patients received hypofractionated IMRT. The cumulative incidences of acute esophagitis grade ≥2 [38.9% (42/108) vs 35.0% (36/103), P=0.895] and grade ≥ 3 [1.9% (2/108) vs 5.8% (6/103), P=0.132] were similar between conventional and hypofractionated IMRT group. Late esophagus injuries grade ≥2 occurred in one patient in either group. No differences in the cumulative incidence of acute pneumonitis grade ≥2[12.0% (13/108) vs 5.8% (6/103), P=0.172] and late lung injuries grade ≥2[5.6% (6/108) vs 10.7% (11/103), P=0.277] were observed. There was no grade ≥3 lung injuries occurred in either group. Using multiple regression analysis, mean esophageal dose ≥13 Gy (OR=3.33, 95% CI: 1.23-9.01, P=0.018) and the overlapping volume between planning target volume (PTV) and esophageal ≥8 cm(3)(OR=3.99, 95% CI: 1.24-12.79, P=0.020) were identified as the independent risk factors associated with acute esophagitis grade ≥2 in the hypofractionated radiotherapy group. Acute pneumonitis grade ≥2 was correlated with presence of chronic obstructive pulmonary disease (COPD, P=0.025). Late lung injuries grade ≥2 was correlated with tumor location(P=0.036). Conclusions: Hypofractionated IMRT are tolerated with manageable toxicities for limited-stage SCLC patients treated with IMRT. Mean esophageal dose and the overlapping volume between PTV and esophageal are independently predictive factors of acute esophagitis grade ≥2, and COPD and tumor location are valuable factors of lung injuries for limited-stage SCLC patients receiving hyofractionated radiotherapy. Prospective studies are needed to confirm these results.
Humans ; Small Cell Lung Carcinoma/pathology* ; Lung Neoplasms/pathology* ; Radiotherapy, Intensity-Modulated/methods* ; Retrospective Studies ; Lung Injury ; Radiotherapy Dosage ; Radiation Injuries/epidemiology* ; Esophagitis/epidemiology* ; Risk Factors ; Pulmonary Disease, Chronic Obstructive/complications*

OBJECTIVETo investigate the clinical features of nutritional iron deficiency anemia (IDA) and analyze the risk factors for the severity of anemia, and to provide a basis for the prevention and clinical diagnosis and treatment of this disease.

METHODSA retrospective analysis was performed on the clinical data of 372 children with IDA to investigate the risk factors for the severity of IDA.

RESULTSOf 372 cases, the male-to-female ratio of these patients was 2.72 : 1. Of all cases, 79.9% were aged 6 months to 2 years, and 30.7% were premature infants; 22.9% had a birth weight of < 2.5 kg, and 77.1% had a birth weight of ≥2.5 kg; 36.0% were delivered by natural birth, and 64.0% were delivered by caesarean section; 79.3% were not given solid foods in time; 46.2% had a history of lower respiratory tract infection/recurrent upper respiratory tract infection, diarrhea, trauma, or surgery. The univariate analysis showed that age, birth weight, gestational age, timely introduction of solid foods, and a history of lower respiratory tract infection/recurrent upper respiratory tract infection, diarrhea, trauma, or surgery were associated with the severity of anemia. The multivariate analysis showed that birth weight and the mentioned medical history were associated with the severity of anemia.

CONCLUSIONSNutritional IDA is common in children aged 6 months to 2 years. Nowadays, improper feeding pattern is still one of the main causes of IDA. Birth weight and a history of lower respiratory tract infection/recurrent upper respiratory tract infection, diarrhea, trauma, or surgery are closely associated with the severity of anemia.

Adolescent ; Anemia, Iron-Deficiency ; etiology ; Child ; Child, Preschool ; Female ; Humans ; Infant ; Logistic Models ; Male ; Retrospective Studies ; Risk Factors

OBJECTIVETo investigate the clinical features of Epstein-Barr virus-related hemophagocytic lymphohistiocytosis (EBV-HLH), to analysis the outcome of HLH-2004 protocol, and to explore the prognostic factors in EBV-HLH patients.

METHODSThe clinical features at onset and outcome of HLH-2004 protocol from 83 pediatric patients with EBV-HLH enrolled from January 2006 to December 2009 in our hospital were analyzed retrospectively. Univariate and multivariate COX regression analysis were used to identify statistically significant prognostic factors.

RESULTS(1) Among the 83 patients, 45 were males and 38 were females. The age of onset ranged from 6 months to 14 years 4 months. 44 patients were treated with HLH-2004, and 3-year overall survival (OS) was (55.8 ± 7.9)%. (2) The most common clinical features of EBV-HLH included high fever, cytopenia, hepatosplenomegaly, and coagulopathy; The respiratory symptoms, angina phlogistic, skin rashes, neurologic abnormality were rare. 97.3% of patients showed an elevation of serum ferritin, liver dysfunction and lipid metabolism disorders was found in most of EBV-HLH patients. 89.0% of patient had hemophagocytosis in bone marrow at diagnosis of EBV-HLH. (3) COX regression analysis revealed that anemia degree, serum albumin < 30 g/L, CD4:CD8 abnormity, NK cell < 3%, treatment protocol were related with the prognosis significantly (P < 0.05).

CONCLUSIONEBV-HLH in pediatric patients has severe clinical feature and poor prognosis. HLH-2004 protocol is an effective treatment for patients with EBV-HLH. Symptomatic treatment can't rescue the patients of EBV-HLH.

Adolescent ; Child ; Child, Preschool ; Epstein-Barr Virus Infections ; drug therapy ; Female ; Herpesvirus 4, Human ; Humans ; Infant ; Killer Cells, Natural ; Lymphohistiocytosis, Hemophagocytic ; drug therapy ; virology ; Male ; Prognosis ; Retrospective Studies ; Treatment Outcome

OBJECTIVETo summarize the clinical characteristics of secondary coagulation disorders caused by exposure to poison (raticide) in children and to investigate the diagnosis and corresponding treatment.

METHODThe process of diagnosis, clinical characteristics, response to treatment and the prognosis were analyzed.

RESULTSThe main clinical manifestation was mucosal bleeding (66.6%), including epistaxis, gingival bleeding, hematomas and so on. All these children were previously well and had no history of bleeding. Activated partial thromboplastin time (APTT) and prothrombin time (PT) were prolonged, factor II was undetectable and the levels of factors VII, IX, and X were lower. The fibrinogen was normal. A raticide was detected in blood and urine of 13 children although 12 of the patients had no definite history of raticide ingestion. Prothrombin complex, fresh frozen plasma and vitamin K(1) were effective in these cases. However, 2 - 3 weeks later, 6 patients presented with recurrent bleeding.

CONCLUSIONFor children with secondary coagulation disorders of unknown cause, intoxication of raticide should be considered. The administration of blood coagulation factors and vitamin K(1) are effective in early treatment, and the treatment period should be more than 2 months. The PT and APTT should be followed up. Vitamin K(1) should be stopped when PT and APTT are normal.

Blood Coagulation Disorders ; chemically induced ; diagnosis ; therapy ; Female ; Humans ; Infant ; Infant, Newborn ; Male ; Rodenticides ; poisoning ; Vitamin K 1 ; administration & dosage ; therapeutic use

OBJECTIVEWiskott-Aldrich syndrome (WAS) is a primary immunodeficiency diseases. The patients with classical WAS have poor prognosis. The hematopoietic stem cell transplantation is the most effective method to cure WAS at present. In this report, a patient with WAS was cured with HLA identical sibling bone marrow transplantation (BMT).

METHODSWiskott-Aldrich syndrome protein (WASP) was detected using flow cytometry and WASP were analyzed for the diagnosis. The bone marrow was collected from the elder sister who was the HLA identical sibling donor. A total of 4.38x10(8)/kg mononuclear cell (MNC) and 3.78x10(6)/kg CD34+ cells were collected and transfused into the patient after the conditioning regimen with busulfan/cyclophosphamide. Cyclosporine only was used for graft-versus-host disease prophylaxis. WASP and short tandem repeats (STR) were detected as the evidence of engraftment.

RESULTSThe diagnosis was WAS: WASP (-IVS9+2T>C, WASP-negative). The patient received busulfan/cyclophosphamide 9 days before the transplantation. WBC decreased to 0.1x10(9)/L in d+4; The absolute number of neutrophils (ANC) was 0.8x10(9)/L in d+13, and exceeded 1.0x10(9)/L later on. From d(-9)-d+14 the patient was dependent on platelet transfusion. From d+15 the patient's PLT>50x10(9)/L and returned to normal after d+30. In d+9-d+10 mild GVHD (I degree) occurred but subsided after the steroid treatment. From d+50, WASP was detected positive and STR showed full donor DNA chimera. Follow-up for 510 d post-transplant, the patient suffered only from mild cold twice, no eczema, no bleeding occurred. The PLT is normal and no chronic GVHD occurred. The levels of IgG, IgM and IgA of the patient were approximately normal.

CONCLUSIONThe HLA-identical sibling's BMT seems to be the periorit treatment of choice for the WAS patient.

Child, Preschool ; Hematopoietic Stem Cell Transplantation ; Humans ; Male ; Treatment Outcome ; Wiskott-Aldrich Syndrome ; surgery

OBJECTIVETo analyze the clinical and laboratory data from acute lymphoblastic leukemia (ALL) patients and the results of treatment using 04 Protocol (suggested by the Pediatric Hematology Group of Chinese Medical Association in 2004).

METHODSThis study included 88 children with ALL below the age of 18 years during the period from October 1, 2004 to June 30, 2007. Minimal inhibitory concentration (MIC) and clinical risk classification were done and the new chemotherapy regimen was used according to the protocol. Patients were stratified into low-risk (LR), medium-risk (MR), and high-risk (HR) groups. Life table method was used to estimate survival rate and statistical analysis was done by using software SPSS for Windows.

RESULTSFrom October 2004 to June 2007, 88 childhood ALL patients were treated with the 04 Protocol. Sixty-three (91.30%) patients attained complete remission (CR) and 17 patients lost to follow up. The overall 4-year-event-free survival (EFS) rate (+/- SE) was (59.73 +/- 7.22)%. EFS was (75.60 +/- 9.71)% in the LR (n = 30), (65.50 +/- 11.69)% in the MR (n = 20) and (44.03 +/- 12.36)% in the HR. Relapse occurred in 18.18% of patients. Seven (7.95%) of 88 patients with ALL died during he induction therapy. Infection was the most common cause of death.

CONCLUSIONThe outcome of patients treated with the 04 Protocol was favorable. Clinical risk classification and the leukemia cells of D19 are independent predictors of prognosis of ALL. High dose methotrexate played an important role in prevention and treatment of central nervous system leukemia. The mortality rate of this chemotherapeutic protocol during induction therapy was high.

Adolescent ; Antineoplastic Combined Chemotherapy Protocols ; therapeutic use ; Child ; Child, Preschool ; China ; Female ; Humans ; Infant ; Male ; Precursor Cell Lymphoblastic Leukemia-Lymphoma ; diagnosis ; drug therapy ; Retrospective Studies ; Risk Factors ; Treatment Outcome

OBJECTIVETo study the potential effect of hypoxia on invasion and metastasis of leukemia cell line K562.

METHODSK562 cells were cultured with the conventional method in vitro and treated with 1%, 3% and 5% oxygen for 24 hrs. The normoxic cultured K562 cells were used as the control group. Cell adhesion assay, cell migration assay and cell invasion assay were used to detect the adhesion, migration and invasion abilities of K562 cells. RT-PCR was used to measure the mRNA expression of HIF-1alpha, VEGF, MMP-2 and MMP-9. The protein level of HIF-1alpha was measured by Western blot.

RESULTSCompared with the control group, the 3% and 5% oxygen treatment groups significantly increased the adhesion, migration and invasion abilities of K562 cells (p<0.05 or <0.01), and up-regulated the protein level of HIF-1alpha and the mRNA levels of HIF-1alpha,VEGF, MMP-9 and MMP-2 (p<0.05 or 0.01). However, there were no significant differences in the above indexes between the 1% oxygen treatment and the control groups.

CONCLUSIONSModerate hypoxia can enhance the abilities of invasion and metastasis of K562 cells, probably by an up-regulation of HIF-1alpha level and VEGF, MMP-2 and MMP-9 mRNA expression.

Animals ; Cell Adhesion ; Cell Hypoxia ; Cell Movement ; Humans ; Hypoxia-Inducible Factor 1, alpha Subunit ; genetics ; K562 Cells ; Matrix Metalloproteinase 2 ; genetics ; Matrix Metalloproteinase 9 ; genetics ; Mice ; NIH 3T3 Cells ; Neoplasm Invasiveness ; Neoplasm Metastasis ; Vascular Endothelial Growth Factor A ; genetics

OBJECTIVETo establish the tridimensional culture method for tissue-engineered skin to observe the histomorphological change in human immortal KC strain (HacaT)cocultured with xenogenic acellular dermal matrix (ADM).

METHODSThe ADM was prepared from SD rats by a modified method. HaCaTs were cultured in defined KC-serum free medium. HaCaTs in log growth phase were inoculated on ADM at the cell density of 2 x 10(5)/cm(2). They were submergedly cultured for 5 days and then changed to air-liquid phase culture for another 5 days. ADM and growth of HaCaTs on day 1 and 5 after cocultured with ADM were observed with scanning electron microscope. The histological change in ADM and HaCaTs on day 1, 5, and 10 after cocultured with ADM were examined by HE staining.

RESULTSThe gross appearance of ADM was white with smooth and soft texture, and intact collagen bundles without cellular residue. HaCaTs adhered and stretched out pseudopodia on the surface of the ADM on day 1 after combined culture, and a monolayer of cells was formed on day 5, growing into 3-6 layers of cells on day 10 with a tendency to grow into ADM.

CONCLUSIONSSD rats ADM is benefit for the adhesion of HaCaTs and the permeation of nutrient solution, from which an engineered multiple-layered human skin can be obtained within 10 days.

Animals ; Cells, Cultured ; Coculture Techniques ; Dermis ; cytology ; Humans ; Keratinocytes ; cytology ; Rats ; Rats, Sprague-Dawley ; Skin, Artificial ; Tissue Engineering ; methods

OBJECTIVETo study the inhibitory effects of matrine, in different concentrations, on invasion and metastasis of human acute lymphocytic leukemia cell line Jurkat.

METHODSIn vitro cultured Jurkat cells were treated by matrine in concentration of 0 g/L, 0.1 g/L, 0.15 g/L and 0.2 g/L, respectively. Then cell adhesion assay, cell migration assay and matrigel invasion assay were used respectively to observe the effects of matrine on adhesion, migration and invasive capacity of Jurkat cells. Meantime, RT-PCR was performed to detect the matrix metalloproteinase (MMP)-2 and MMP-9 mRNA expression levels. Comparison of measurement data among groups was analyzed by variance analysis.

RESULTSAs compared with the control group, the adhesion of Jurkat to fibronectin (FN) was significantly inhibited by 0.15 g/L and 0.2 g/L of matrine (P < 0.05); the cell migration and invasive capacity were significantly lowered by 0.1 g/L, 0.15 g/L and 0.2 g/L matrine (P < 0.01). High mRNA expression of MMP-9 presented but that of MMP-2 was expressed insignificantly in Jurkat cells, matrine at 0.1 g/L, 0.15 g/L and 0.2 g/L showed obvious effect in down-regulating MMP-9 mRNA expression (P < 0.01). Besides, MMP-9 mRNA expression was found to be positively correlated with the invasive capacity of Jurkat cells (r = 0.940, P < 0.01).

CONCLUSIONSMatrine is a good drug for antagonizing the invasion and metastasis of leukemia cells, it may roundly inhibit the adhesion, migration and invasive capacity of Jurkat cells, the mechanism might be related with the down-regulation of MMP-9 mRNA expression.

Alkaloids ; pharmacology ; Animals ; Cell Proliferation ; drug effects ; Down-Regulation ; Drugs, Chinese Herbal ; pharmacology ; Humans ; Jurkat Cells ; Leukemia ; drug therapy ; pathology ; physiopathology ; Mice ; NIH 3T3 Cells ; Neoplasm Invasiveness ; Neoplasm Metastasis ; Quinolizines ; pharmacology ; Random Allocation