中国实验血液学杂志最新文献

Myeloproliferative neoplasms (MPN) are a group of malignant myeloid tumors caused by hematopoietic stem cell proliferation. The discovery of gene mutations has elucidated the pathogenesis of MPN and provided molecular diagnostic criteria for MPN. Recent studies have shown that there are cytokine disorders in MPN patients, and the changes in the microenvironment caused by these cytokine disorders may have great significance for the pathophysiology and pathogenesis of MPN, which may lead to corresponding clinical symptoms and different prognosis in patients. In this review, the latest research progress on the role and status of cytokines in MPN will be summarized.

Objective: To analyze and explore the effects of Huayu Jiedu Decoction on the malignant biological characteristics of multiple myeloma (MM) cells and its molecular mechanism, so as to provide experimental basis and theoretical basis for the alternative therapy of anti-MM in traditional Chinese medicine.

Methods: Different concentrations of Huayu Jiedu Decoction were used to intervene myeloma U266 cells. The changes of cell proliferation activity were detected by CCK-8 assay, apoptosis was detected by Annexin V/PI double staining flow cytometry, and apoptosis and protein expression of related signaling pathways were detected by Western blot. Real-time quantitative PCR was used to detect mRNA expression changes of high mobility group protein B1 (HMGB1), CXC chemokine receptor 4 (CXCR4) and interleukin-6 (IL-6).

Results: Huayu Jiedu Decoction inhibited the proliferative activity of U266 cells and induced their apoptosis in a concentration and time dependent manner ( r =-0.713, r =-0.827). After treatment with Huayu Jiedu Decoction for 48 h, the expressions of anti-apoptotic protein Bcl-2 and survivin were down-regulated, while the expression of pro-apoptotic protein Bax was up-regulated, and the phosphorylation level of TLR4/NF-κB signaling pathway was inhibited. After intervention of Huayu Jiedu decoction, the expressions of HMGB1 and IL-6 mRNA were significantly decreased, while the expression of CXCR4 was not significantly decreased.

Conclusion: Huayu Jiedu Decoction can inhibit the proliferative activity of U266 cells and induce programmed death. Its molecular mechanism may be related to regulating the expression of apoptotic proteins, inhibiting the activation of TLR4/NF-κB pathway and down-regulating the expression of HMGB1 and IL-6 mRNA.

Objective: To investigate how reactive oxygen species (ROS) regulates the signal transduction of platelet activation and apoptosis, and to explore the relationship between platelet activation and apoptosis.

Methods: Platelets were directly stimulated with thrombin or pretreated with ROS inhibitor N-acetylcysteine (NAC) before being stimulated with thrombin, and then flow cytometry was used to detect the effects of thrombin and NAC on P-selectin expression, αⅡbβ3 activation, mitochondrial membrane potential depolarization, phosphatidylserine (PS) externalization, ROS expression and platelet aggregation.

Results: Thrombin could induce the production of ROS in platelets in a concentration- and time-dependent manner. 0.01 U thrombin induced ROS-dependent high degree of integrin αⅡbβ3 activation, P-selectin expression, and platelet aggregation. The platelets induced by different concentration gradients of thrombin exhibited ROS-dependent mitochondrial membrane potential depolarization and PS externalization in platelets. After induction with thrombin for 30 min, the activation of integrin αⅡbβ3 in platelets reached its maximum level, and after 60 minutes, the depolarization of mitochondrial membrane potential in platelets reached its maximum level. However, the expression of P-selectin, depolarization of mitochondrial membrane potential, and platelet aggregation function were all inhibited to a certain extent when the platelets were pretreated with ROS inhibitor NAC and then induced with thrombin.

Conclusion: When platelets are induced by thrombin, ROS first regulates the activation of platelets, and then regulates the apoptosis of platelets. Both platelet activation and apoptosis depend on the production of ROS in platelets, and the signals of activation and apoptosis occur orderly. Inhibiting the ROS signal in platelets can effectively inhibit the activation and apoptosis of platelets.

Objective: To investigate the effects of ATG5 and ATG7 genes on the sensitivity of multiple myeloma (MM) cell line RPMI-8226 cells to ferroptosis.

Methods: CRISPR/Cas9 technology was used to knock out the autophagy key genes ATG5 and ATG7 in RPMI-8226 cells. Western blot was used to identify gene knockout cells, and detect the expression changes of autophagy-related proteins P62 and LC3B. Flow cytometry was used to detect the change of sensitivity of gene knockout cells to RSL3. The content of intracellular ferrous ions and reactive oxygen species (ROS) level in gene knockout cells were detected.

Results: Western blot result confirmed that ATG5 and ATG7 genes were knocked out successfully in RPMI-8226 cells. The result of flow cytometry showed that the cell viability of RPMI-8226 cells was dose-dependent on different concentrations of RSL3 (r =-0.969). RSL3 (10 μmol/L) was used to induce ferroptosis in cells of control group and gene knockout groups, then the cell viability in gene knockout groups were both higher than control group after 48 hours (both P < 0.001). After knocking out the ATG5 and ATG7 genes, the content of intracellular Fe²⁺ decreased significantly compared with control group (both P < 0.01), and the ROS level also decreased (both P < 0.001).

Conclusion: Knockout of ATG5 and ATG7 genes can inhibit the ferroptosis of MM cells, and LAP pathway may be involved in the regulation.

Objective: To retrospectively analyze the detection and diagnosis process of two cases with double rare thalassemia genotypes, explore the causes of missed diagnosis and misdiagnosis of rare thalassemia, and improve the diagnosis level of rare thalassemia.

Methods: Base on the family history, hematological phenotype and hemoglobin electrophoretic analysis results, the common genotypes of α and β-thalassemia were detected by PCR+diversion hybridization. DNA sequencing technology was used for rare α and β protein genes sequencing.

Results: Both subjects were combined with double rare thalassemia genotypes, and both rare thalassemia gene combinations were reported for the first time. One of them was αβ complex thalassemia with αα^{*53_55 del TCC}/αα heterozygous merger β^{IVS II-2(-T)}/β^N heterozygous, the other was αα^{IVS-II-55(T→G) in α1}/αα^4.2-Q double azygous heterozygous α-thalassemia, among which αα^{*53_55 del TCC}/αα genotype was also reported for the first time.

Conclusion: The reported rare gene type αα^{*53_55 del TCC}/αα and two cases of rare gene combinations enriches the spectrum of gene mutations in the Chinese population, and provides richer molecular information for thalassemia diagnosis and eugenics counseling.

Objective: To establish a highly sensitive and quantitative detection method for ABL1 kinase region mutations, provide strong support for the early diagnosis and treatment of chronic myeloid leukemia(CML).

Methods: Sampele from 35 CML patients who were initially tested negative for ABL1 kinase region mutations by Sanger sequencing were collected. The ABL1 kinase region mutation was detected by the fluorescence quantitative detection kit of Shanghai Yuanqi Biopharmaceutical Technology Co., Ltd. The mutation rate was analyzed by ^△△Ct value method. The relative mutation rate of the final ABL1 kinase region was determined by dividing the mutation rate by the expression level of the fusion gene.

Results: Among the 35 CML patients initially tested negative for ABL1 mutations by the Sanger sequencing method, 7 cases of T315I mutation, 2 cases of T315A mutation, 2 cases of Y253H mutation, and 1 cases of E255K mutation after detection of the new method. The relative mutation rates range from 0.1% to 19.42%, which could not be detected by Sanger sequencing method. Subsequently, this method was used to detect the ABL1 mutation in 126 CML patients, and the positive rate exceeded that of the Sanger sequencing method. The BCR-ABL1 gene expression significantly reduced or negative after adjusting treatment strategy based on the mutation situation.

Conclusion: Compared with Sanger sequencing, fluorescence quantitative PCR has higher sensitivity and can screen for low-frequency ABL1 kinase mutations in the early stage. Moreover, it can also perform relative quantitative analysis, so the method has good clinical application prospects for detecting ABL1 mutation.

Objective: To detect the pharmacokinetic (PK) parameters of coagulation factor Ⅷ (FⅧ) in adult patients with severe hemophilia A, identify the potential factors influencing FⅧ PK, and optimize the use of FⅧ in individual prophylaxis regimens.

Methods: PK characteristics of FⅧ were studied in a total of 23 severe hemophilia A adults. The correlation of patients' characteristics including age, von Willebrand factor antigen (vWF:Ag), blood group, weight, body mass index (BMI) and FⅧ genotype, with FⅧ PK were evaluated. Individual prophylaxis regimens were given based on FⅧ PK parameters.

Results: The mean terminal half‑life (t_1/2) of FⅧ was 20.6±9.3 h, ranged from 11.47 h to 30.12 h. The age (r =0.580) and vWF:Ag (r =0.814) were significantly positively correlated with t_1/2 of FⅧ. The mean area under the plasma concentration curve (AUC) of FⅧ was 913±399 (328-1 878) IU·h/dl, and the AUC of FⅧ was positively correlated with age (r =0.557) and vWF:Ag (r =0.784). The mean residence time (MRT) of FⅧ was 24.7±12.4 (13.2-62.2) h, and the MRT of FⅧ was positively correlated with age (r =0.664) and vWF:Ag (r =0.868). The mean in vivo recovery (IVR) of FⅧ was 2.59±0.888 (1.5-4.29) IU/dl per IU/kg, the mean clearance (CL) of FⅧ was 3±1.58（0.97-7.18）ml/(kg·h)，and there was no significant correlation of IVR and CL with age and vWF:Ag. According to the individual PK parameters, ultra low-dose, low-dose and moderate-dose FⅧ were applied to 15, 6, 2 adults patients with severe hemophilia A for prophylaxis, respectively.

Conclusion: There are significant individual differences in the FⅧ half-life of adult patients with severe hemophilia A. The older the patient, the higher the vWF:Ag level, and the longer the FⅧ half-life. Individual administration is required based on the FⅧ PK parameters to optimize prophylaxis treatment.

Objective: To investigate the efficacy and safety of ixazomib combined with thalidomide and dexamethasone in the treatment of multiple myeloma (MM).

Methods: The clinical data of 60 MM patients admitted to our center from January 2019 to June 2022 were analyzed retrospectively, including 43 newly diagnosed patients and 17 patients with recurrence and progression. All patients were treated with ixazomib combined with thalidomide and dexamethasone, and completed 2 to 7 treatment cycles.

Results: The overall response rate (ORR) of all patients was 98.3%. Among them, 53 patients completed 4 treatment cycles, and the ORR was 86.8%. Seventeen patients completed the whole treatment cycle, with curative effect reaching 88.2% achieving very good partial response and above, and 52.9% achieving complete response and above. Albumin and β₂-microglobulin of all patients had been improved rapidly after treatment. The deadline was August 31, 2022. The median follow-up time was 14(3-24) months, and overall survival (OS) rate was 86.67%. The OS rate of patients with recurrence and progression was significantly lower than that of newly diagnosed patients (P < 0.05). The most common adverse reaction of hematology was lymphopenia (53.3%), followed by anemia (33.3%). The most common non-hematological adverse reaction was fatigue (68.33%), followed by peripheral neuropathy (31.67%).

Conclusion: Ixazomib combined with thalidomide and dexamethasone is effective in the treatment of MM, with good short-term efficacy, survival and safety. However, its long-term efficacy needs further observation.

Objective: To explore the clinical application value of the Bleeding Scoring Scales (2019 Pediatric ITP Scale) in the diagnosis and treatment of children with primary immune thrombocytopenia (ITP).

Methods: A total of 422 children with ITP were evaluated with the 2019 Pediatric ITP Scale and the 2013 ITP-BAT and their clinical data were analyzed. The correlation between the two bleeding scoring scales and disease stage, platelet count was analysed, the evaluation time, consistency of the two bleeding scoring scales was compared, and the correlation of the two methods. The changes of platelet count and the score of 2019 Pediatric ITP Scale before treatment and after treatment at 48 h and one week were analyzed to detect responsiveness of the 2019 Pediatric ITP Scale.

Results: The score of the 2019 Pediatric ITP Scale was mainly one point and two points, the corresponding bleeding was skin and mucosal bleeding.404 patients (95.7%) had bleeding manifestations, including 249 patients of skin bleeding (59.0%), 144 patients of mucosal bleeding (34.1%), and 11 patients of organ bleeding (2.6%), of which 2 patients were severe bleeding. The two bleeding scoring scales were both negatively correlated with platelet counts in children with ITP (r _s=-0.5032, r _s=-0.6084) and no correlation with the stage of pediatric ITP(P ＞0.05). The 2019 Pediatric ITP Scale had good consistency with the 2013 ITP-BAT (r _s=0.7638). The average time required for the 2019 Pediatric ITP Scale was 88.64 (40-181) seconds, which was lower than that required for the 2013 ITP-BAT 104.12 (47-285) seconds (Z =17.792, P < 0.001). The 2019 Pediatric ITP Scale can well reflect the treatment of pediatric ITP. There were statistically significant differences in platelet count before treatment and after treatment at 48 h and one week among steroid group, IVIG group and steroid combined with IVIG group ( P < 0.05). There were also statistically significant differences in the score of the 2019 Pediatric ITP Scale before treatment and after treatment at one week among the three groups ( P < 0.05).

Conclusion: The 2019 Pediatric ITP Scale has good consistency and sensitivity in clinical application, and it takes less time to complete than the 2013 ITP-BAT, this scale can be used as an effective tool for disease assessment and efficacy determination in pediatric primary immune thrombocytopenia.

Research on the participation of exosomes in pathogenesis and prognosis of multiple myeloma (MM) has made encouraging progress. However, the specific mechanism has not yet been clarified. Recently, domestic and foreign researchers have pressed forward on deeper study on exosomes. This article mainly summarizes the role of exosomes in the pathophysiological processes, such as bone marrow microenvironment changes, immunosuppression, myeloma bone disease generation and myeloma drug resistance, so as to provide new reference for further clinical research of exosomes as potential biomarkers for MM.