OBJECTIVE: To investigate the efficacy, prognosis and safety of decitabine combined with modified EIAG regimen in the treatment of patients with relapsed/refractory acute myeloid leukemia (AML) and high-risk myelodysplastic syndrome (MDS). METHODS: The clinical data of 44 patients with relapsed/refractory AML and high-risk MDS admitted to our hospital from January 2017 to December 2020 were analyzed retrospectively. The patients were equally divided into D-EIAG group (decitabine combined with EIAG regimen) and D-CAG group (decitabine combined with CAG regimen) according to clinical treatment regimen. The complete response (CR), CR with incomplete hematologic recover (CRi), morphologic leukemia-free state (MLFS), partial response (PR), overall response rate (ORR), modified composite complete response (mCRc), overall survival (OS) time, 1-year OS rate, myelosuppression and adverse reactions between the two groups were compared. RESULTS: In D-EIAG group, 16 patients (72.7%) achieved mCRc (CR+CRi+MLFS), 3 patients (13.6%) achieved PR, and ORR (mCRc+PR) was 86.4%. In D-CAG group, 9 patients (40.9%) achieved mCRc, 6 patients (27.3%) achieved PR, and ORR was 68.2%. Difference was observed in mCRc rate between the two groups (P=0.035), but not in ORR (P>0.05). The median OS time of D-EIAG group and D-CAG group was 20 (2-38) months and 16 (3-32) months, and 1-year OS rate was 72.7% and 59.1%, respectively. There was no significant difference in 1-year OS rate between the two groups (P>0.05). After induction chemotherapy, the median time for absolute neutrophil count recovery to 0.5×10⁹/L in D-EIAG group and D-CAG group was 14 (10-27) d and 12 (10-26) d, for platelet count recovery to 20×10⁹/L was 15 (11-28) d and 14 (11-24)d, the median red blood cell suspension transfusion volume was 8 (6-12) U and 6 (6-12) U, and the median apheresis platelet transfusion volume was 4 (2-8) U and 3 (2-6) U, respectively. There were no statistically significant differences in comparison of the above indicators between the two groups (P>0.05). The hematological adverse reactions of patients were mainly myelosuppression. Grade III-IV hematological adverse events occurred in both groups (100%), with no increase in the incidence of non-hematological toxicities such as gastrointestinal reactions or liver function damage. CONCLUSION Decitabine combined with EIAG regimen in the treatment of relapsed/refractory AML and high-risk MDS can improve remission rate, provide an opportunity for subsequent therapies, and have no increase in adverse reactions compared with D-CAG regimen.
Humans ; Decitabine/therapeutic use* ; Treatment Outcome ; Retrospective Studies ; Cytarabine ; Myelodysplastic Syndromes/drug therapy* ; Leukemia, Myeloid, Acute/drug therapy* ; Bone Marrow Diseases/drug therapy* ; Antineoplastic Combined Chemotherapy Protocols/therapeutic use*

OBJECTIVE: To explore the molecular biology of D variant blood group with RHD*95A genotype and the genetic mechanism of its generation. METHODS: A total of 6 samples from 3 generations of a family were analyzed. RHD blood group was identified by saline test tube and microcolumn gel card method. 10 exons of RHD gene were amplified by Polymerase Chain Reaction-Sequence Specific Primer (PCR-SSP) and analyzed by direct sequencing. Homology modeling was used to compare the structural differences between mutant RHD protein and wild-type RHD protein. RESULTS: The proband was identified as D variant by serological identification, RHD gene sequencing directly detected a c. 95 c > A mutation in exon 1 that leads to encoding the 32-bit amino acids by threonine Thr (T) into aspartic acid Asn (N), the rest of the exon sequences were normal compared with the normal RHD*01 gene. In the family, the proband's father, grandmather and uncle were all carried the same RHD*95A allele. Protein modeling results suggested that the hydrogen chain connected to the 32nd amino acid residue was changed after p.T32N mutation, which affected the structural stability of RHD protein. CONCLUSION The first genetic lineage of the RHD*95A gene was identified in a Chinese population. The c.95C>A mutation in RHD gene was found in the family, which resulted in reduced expression of RHD antigen and showed D variant, the mutation could be stably inheritable. Gene identification and protein structure analysis of D variant population is helpful to explore the molecular mechanism of its formation and ensure the safety of blood transfusion.
Humans ; Blood Group Antigens

OBJECTIVE: To study the effect and safety of G-CSF combined with Plerixafor on the mobilization of peripheral blood hematopoietic stem cells from healthy related donors of allogeneic hematopoietic stem cell transplantation (allo-HSCT). METHODS: It was analyzed retrospectively that the data of peripheral blood hematopoietic stem cells from 33 (observation group) related donors mobilized by G-CSF plus Plerixafor in Hebei Yanda Lu Daopei Hospital from April 2019 to April 2021. Bone marrow and peripheral blood hematopoietic stem cells (PBSCs) of these donors were respectively collected on the fourth and fifth day of G-CSF-induced mobilization. Following the administration of Plerixafor on the night of the fifth day, PBSCs were collected on the sixth day once again. 46 donors using "G-CSF only" mobilization method in the same period were randomly selected as the control and respectively analyzed the differences of CD34+ cell counts on the fifth and the sixth day in two groups. And the donors' adverse reaction to Plerixafor in the form of questionnaire was also observed. Then it was compared that the patients who underwent allo-HSCT in "G-CSF+Plerixafor" group and "G-CSF only" group in terms of acute GVHD at grade I-IV or III-IV, CMV reactivation and EBV reactivation. RESULTS: CD34+ cells count (M±Q) among PBSCs collected on the fifth and the sixth day in the observation group were (1.71±1.02)×10⁶/kg and (4.23±2.33)×10⁶/kg, respectively. CD34+ cell counts on the sixth day was significantly higher than that of the fifth day (P<0.001); While the counterparts in the control group were (2.47±1.60)×10⁶/kg and (1.87±1.37)×10⁶/kg, respectively. By statistical analysis, CD34+ cell counts on the sixth day was significantly less than that of the fifth day (P<0.001). The adverse reaction to Plerixafor for the donors in the study were all grade 1 or 2 (mild or moderate) according to CTCAE 5.0 and disappeared in a short time. The patients who underwent allo-HSCT in the "G-CSF+Plerixafor" group and "G-CSF only" group were not statistically significant in terms of acute GVHD at grade I-IV or III-IV, CMV reactivation and EBV reactivation (P>0.1). CONCLUSION The cell mobilization program of G-CSF combined with Plerixafor is safe and effective for being applied to allo-HSCT. The addition of Plerixafor can significantly increase the number of CD34 postive cells in the PBSC collection. Key words　　; ;
Antigens, CD34 ; Benzylamines ; Cyclams ; Granulocyte Colony-Stimulating Factor ; Hematopoietic Stem Cell Mobilization ; Hematopoietic Stem Cell Transplantation ; Hematopoietic Stem Cells ; Heterocyclic Compounds ; Humans ; Peripheral Blood Stem Cell Transplantation ; Retrospective Studies

OBJECTIVE: To analyze the clinical characteristics of bloodstream infection (BSI) in patients treated by hematopoietic stem cell transplantation (HSCT). METHODS: The clinical characteristics, distribution of pathogenic bacteria causing BSI and drug sensitivity of 910 patients treated by HSCT in our department from January 2013 to June 2020 were retrospectively analyzed. RESULTS: Among 910 HSCT patients, 111 patients were diagnosed as BSI within 100 days after transplantation, and 98 patients showed BSI during the period of agranulocytosis. Multivariate analysis showed that the usage of anti-thymocyte globulin (ATG), long duration of agranulocytosis and low infusion volume of mononuclear cell (MNC) were the independent risk factors affecting BSI after HSCT. Among 121 pathogenic bacteria isolated, 76 Gram-negative (G^-) bacteria (62.8%), 40 Gram-positive (G⁺) bacteria (33.0%), and 5 fungi (4.1%) were detected out. The top three pathogens were Escherichia coli, Staphylococcus epidermidis and Pseudomonas aeruginosa. The drug-resistance rates of Escherichia coli and Klebsiella pneumoniae to carbapenems was 14.3% and 7.7%, respectively, and Pseudomonas aeruginosa was 66.7%. The susceptibility of G⁺ bacteria to vancomycin, linezolid and teicoplanin was 97.5%, 100% and 100%, respectively. The crude mortality rate of the patients with BSI at 100 days after HSCT was significantly higher than that of patients without BSI (P<0.001). CONCLUSION The usage of ATG, long duration of agranulocytosis and low infusion volume of MNC are independent risk factors for BSI after HSCT. The pathogens after HSCT are mainly G- bacteria. Pseudomonas aeruginosa is highly resistant to carbapenems. Key words　　;
Bacteremia/epidemiology* ; Bacteria ; Hematopoietic Stem Cell Transplantation ; Humans ; Retrospective Studies ; Sepsis

OBJECTIVE: To analyze the proliferation potential of bone marrow-derived mesenchymal stem cells (MSC) in patients with myelodysplastic syndrome (MDS). METHODS: The MSC derived from the 24 patients with newly diagnosed MDS (MDS-MSC group) and MSC derived from 15 patients with nutritional anemia (control group) in the Affiliated Hospital of Hebei University were used as the research objects. The proliferation potential of MSC was analyzed by colony-forming unit assay, doubling time, cumulative passaging, cell number after 10 days of culture with equal amount of MSC and MTT experiment. The mechanism of abnormal proliferation was analyzed by cell cycle experiment, apoptosis experiment and p21 gene expression assay. RESULTS: In the colony forming unit assay, the number of MDS-MSC colonies was 4.44±2.51, which was significantly lower than that of the control group (12.44±2.55)(P<0.01); the doubling time of MDS-MSC group was significantly longer than that of the control group (7.80±3.26 vs 3.63±0.85) (P<0.01); the number of MDS-MSC in 5×10 CONCLUSION The proliferative capability of MDS-MSC is significantly reduced, which relates with the arrest of cell cycle in G
Apoptosis ; Bone Marrow Cells ; Cell Proliferation ; Humans ; Mesenchymal Stem Cells ; Myelodysplastic Syndromes

Purpose To investigate the expression of insulin-like growth factor Ⅱ mRNA-binding protein 3 (IMP3) and its differential diagnostic significance in benign and malignant thyroid tumor. Methods The fluorescent quantitative PCR and immunohistochemical staining were used to detect the IMP3 expression in 71 cases of thyroid tissue of different pathological types. The differential diagnostic significance of IMP3 expression in benign and malignant thyroid tumor was analyzed. Results Compared with normal thyroid tissue, thyroid tumors including follicular variant of papillary thyroid carcinoma (FVPTC ), follicular thyroid carcinoma (FTC), papillary thyroid carcinoma(PTC), nodular goiter (NG), and follicular adenoma (FA) had significantly higher IMP3 mRNA expression levels with10.13, 8.81, 8.52, 2.46, and 1.49 holds, respectively. The positive expression rate of IMP3 protein in thyroid tumors were significantly higher, with the positive rate from high to low was FTC (100％ ), PTC (96.77％ ), FVPTC (90％ ), FA(20％ ), and NG (0). The expression level of IMP3 protein was positively correlated with the expression of mRNA (P＜0.01). The IMP3 expression level of malignant thyroid tumor(8.82 holds) was significantly higher than that of benign thyroid tumor (1.94 holds) (P＜0.01). The IMP3 expression level of malignant thyroid follicular lesions (9.36 holds) was higher than that of benign thyroid follicular lesions (1.49 holds) (P＜0.01). Conclusion IMP3 may be an effective and useful molecular maker for diagnosis of benign and malignant thyroid neoplasms, as well as the differential diagnosis between benign and malignant thyroid follicular lesions.

OBJECTIVETo investigate the blood test patterns for blood donors after nucleic acid detection in blood center.

METHODSThe collected blood samples after voluntary blood donors first were detected by conventional ELISA, then 31981 negative samples were detected via HBV/HCV/HIV combined nucleic acid test of 6 mixed samples(22716 cases) or single samples(9265 cases) by means of Roche cobas s201 instrument. The combined detection method as follows: the blood samples were assayed by conventional nucleic acid test of 6 mixed samples, at same time, 6 mixed samples were treated with polyethylene glycol precipitation method to concentrate the virus, then the nucleic acid test of blood samples was performed; the single detection method as follows: firstly the conventional nucleic acid test of single sample was performed, then the positive reactive samples after re-examination were 6-fold diluted to simulate the nucleic acid test of 6-mixed samples. The positive rate of positive samples detected by combined nucleic acid test, positive samples detected by nucleic acid test of mixed virus concentration and positive samples detected by single nucleic acid test was statistically analyzed. In addition, for HBVpersons the serological test yet should be performed.

RESULTSIn 22 716 samples detected by nucleic acid test of 6 mixed samples (MP-6-NAT) , 9 cases were HBV(0.40‰, 9/22716); at same time, the detection of same samples by nucleic acid test of mixed sample virus concentration showed 29 cases of HBV(1.28‰, 29/22716). In 9265 samples detected by single nucleic acid test(ID-NAT) 12 cases showed HBV(1.30‰, 12/9265), meanwhile the detection of these 12 samples with HBVby 6-fold dilution for virus concentration found only 4 samples with HBV. In serological qualified samples, ID-NAT unqualified rate was 1.28‰, which was higher than that of MP-6-NAT(0.4‰) (χ=8.11, P<0.05); but there was no statistical difference between unqualified rate of ID-NAT and MP-6-NAT(1.3‰ vs 1.28‰)(χ=0.00, P>0.05). In 41 samples with HBsAgHBV DNAdetected by ELISA, 36 samples were confirmed to be occult HBV infective(OBI) by HBsAb, HBcAb test of ELISA; out of these 41 samples, 33 samples showed HBcAb(91.66% of OBI), 5 might be HBV "window period" infective, moreover the HCV RNA and HIV RNA positive samples were not found.

CONCLUSIONTo avoid the missdiagnosis of donors with low level of virus, the nucleic acid test must be carried out after virus concentration of mixed samples when the blood test pattern of donors is nucleic acid test of mixed samples, otherwise the single nucleic acid test must be performed to obtain more high detected rate of virus nucleic acid. The HBcAb serologic test and physical examination of donors before blood donation must be enhanced on basis of serological test of HBsAg; for high risk people, the persuading no blood donation is simplest pattern.

OBJECTIVETo analyze and evaluate the application of spinning disk confocal microscopy and imaging analysis software in movement and phagocytosis of neutrophils.

METHODSNeutrophils were isolated from bone marrow by centrifugation on discontinuous Percoll gradient, and then were stained with PE Gr-1 antibody and mixed with FITC-labeled Zymosan A bioparticles. Multichannel time-lapse videos were captured by using the spinning disk confocal microscopy. The result was analyzed by using volocity and ImageJ software, the parameters associated with movement and phagocytosis of neutrophils were analyzed, including morphological changes, cell tracking, pseudopod dynamics, binding and phagocytosis index.

RESULTSMost neutrophils would be polarized in response to Zymosan particles during a short time. Binding and phagocytosis process occured in forty minutes.

CONCLUSIONA method of precisely quantifying the movement and phagocytosis of neutrophils using microscopic imaging and imaging analysis technique has been set up successfully. Using this method, biological activity and function of neutrophils can be evaluated visually and rapidly. The physiologically rapid response to Zymosan particles can be applied to the neutrophils function research in the future.

Antibodies ; Bone Marrow ; Cell Movement ; Humans ; Microscopy ; Neutrophils ; Phagocytosis ; Zymosan

OBJECTIVETo evaluate the incidence rate of IDH1 in acute myeloid leukemia and analyze its effect on clinical characteristics and prognosis.

METHODSMononuclear cells in bone marrow samples were collected from 192 adult patients with newly diagnosed AML. Polymerase chain reaction (PCR) and direct sequencing were used to amplify exon 4 of IDH1 gene, the gene sequencing was used to analyze the gene mutations, at same time, the detection of NPM1, FLT3-TKD, FLT3-ITD, C-KIT, CEPBA, TET2 and JAK2V617F and MLL mutations were carried out, the follow-up was used to determine its therapeutic efficacy and outcomes of patients. The clinical and laboratory data of these cases were collected, and their clinical characteristics and prognosis were then analyzed.

RESULTSAmong the 192 AML patients, 13 cases were detected with IDH1 gene mutation, the mutation rate was 6.77% [95% CI (5.70%-13.38%)]. The sequencing chart of IDH1 gene showed double peaks, the mutations were heterozygous, out of them c.G395A (p.R132H) was found in 8 cases, c.C394T was found in 4 cases (p.R132C), c.C394A (p.R132S) was found in 1 cases, R132H and R132C are common, 13 cases showed missense mutation. The median age in mutation group was 52 years old, the median age in unnutration group was 40 years, there was significant difference between them (P = 0.010). Mutation rate of IDH1 gene in M1 and M2 was significantly higher than that in other FAB subtypes. There were no significant difference in sex, newly diagnosed peripheral white blood cell count, hemoglobin, platelet count, peripheral blood and bone marrow original cell proportion of primitive cells between them. Mutation of IDH1 gene had certain correlation with NPM1 gene mutation, but no correlation with FLT3-TKD, FLT3-ITD, C-KIT, TET2 and JAK2V617F and MLL natations was found. In addition, the IDH1 mutation easily occurred in patients with normal karyotype or in patients with middle prognostic risk karyotype, IDH1 mutation occurred in 11 cases with normal karyotype, the mutation rate was 10.28%, IDH1 mutation were observed in 2 cases with abnormal karyotype, the mutation rate was 3.50%, there was significant difference. In AML patients with middle prognostic risk karyotype. The complete remission (CR) and the 3 year survival (OS) rate of IDH1 mut patients were less than that in IDH1 wt, there was significant difference (P < 0.05).

CONCLUSIONSThe IDH1 mutation more easily occurr in older AML patients and mutations effect of IDH1 on clinical characteristics may represent a molecular marker for poor prognosis in AML.

Abnormal Karyotype ; Adult ; Exons ; Heterozygote ; Humans ; Isocitrate Dehydrogenase ; metabolism ; Leukemia, Myeloid, Acute ; enzymology ; Leukocyte Count ; Mutation ; Mutation, Missense ; Platelet Count ; Polymerase Chain Reaction ; Prognosis ; Remission Induction ; Survival Rate

This study was aimed to compare the differential expressions of calcineurin (PP2B, PP3) in the mouse Pre-B cell lines (S9) and the tumor cell lines (S4C2) derived from pre-B lymphocytes, and to clarify its possible mechanism involving in the leukemia cell apoptosis. The quantitative real-time PCR was used to detect the differential expressions of H2AX-associated phosphakinase ATM, ATR, DNA-PKs, JNK1, P38 and the γ-H2AX-related phosphatase PP1, PP2A, calcineurin, PP4, PP6, PP5 between S9 and S4C2 cell lines. CCK-8 assay and flow cytometry were used to detect the effect of imatinib (IM) and cyclosporine A (CsA) on cytotoxicity and apoptosis of 2 cell lines. The Western blot was used to detect the effects of 2 drugs on apoptosis of S9 and S4C2 cell lines. The results showed that the expression level of calcineurin gene in the leukemia cell S4C2 was about 3.5 times of that in S9 cells, while the expression of other genes in these 2 kinds of cells was not significantly different. The apoptosis and toxicity of IM and CsA on S4C2 cells was significantly stronger than that on S9 cells. The expression level of calcineurin in S4C2 cells was higher than that in S9 cells.When CsA inhibited the calcineurin activity, the expression of DNA damage marker γ-H2AX in S9 cells was significantly lower than that in S4C2 cells,while the expression level of γ-H2AX between the two cell lines was no significantly different after treatment with imatinib, the expression level of γ-H2AX in S9 cells was lower than that in S4C2 cells when the two drugs were combined. It is concluded that the calcineurin plays a role of anti-apoptosis in B leukemic cells, cyclosporine A can promote the leukemia cell apoptosis.
Animals ; Apoptosis ; Calcineurin ; metabolism ; Cell Line, Tumor ; Cyclosporine ; DNA Damage ; Flow Cytometry ; Leukemia ; metabolism ; Mice ; Precursor Cells, B-Lymphoid ; metabolism ; Real-Time Polymerase Chain Reaction