Bone Marrow Hyperplasia Research Articles

Clinical characteristics of megaloblastic anemia with pancytopenia

ABSTRACT Objectives In this study, we aimed to explore the clinical characteristics of patients with megaloblastic anemia and pancytopenia. Methods Data on patient characteristics, laboratory examinations, clinical manifestations, and associated diseases were collected from Changzhou Hospital of Traditional Chinese Medicine. The participants were divided into two groups according to routine blood test results: megaloblastic anemia patients with pancytopenia (Group A) and megaloblastic anemia patients with simple anemia (Group B). Results The number of patients with fever and bleeding in Group A was much higher than that in Group B. Inpatient days and hospitalization costs in Group A were higher than those in Group B. White blood cell count, red blood cell count, haemoglobin level, and platelet count in Group A were lower than those in Group B. There were no significant differences in mean corpuscular volume (MCV), mean corpuscular haemoglobin (MCH), and serum levels of lactate dehydrogenase (LDH), total bilirubin (TBIL), direct bilirubin (DBIL), indirect bilirubin (IBIL), albumin (ALB), vitamin B12, folic acid, and ferritin between the two groups. The incidence of associated diseases, particularly pulmonary infections, was significantly higher in Group A than in Group B. There was no significant difference in the degree of bone marrow hyperplasia between the two groups. Conclusion Patients with pancytopenia and megaloblastic anemia often developed infection and bleeding, which caused more severe anemia, higher hospitalization costs, and longer inpatient days. Early diagnosis and active treatment are required to reduce patient complications and costs.

Delayed diagnosis of TAFRO syndrome: A case report.

TAFRO syndrome is a systemic inflammatory disorder, manifesting as thrombocytopenia (t), anasarca (a), fever (f), reticulin myelofibrosis/renal insufficiency (r), and organomegaly (o), and considered as a unique clinical subtype of idiopathic multicentric Castleman disease (iMCD). Such syndrome gave rise to a clinical picture similar to that of either a connective tissue disease or an autoimmune disease. A Chinese young female initially presenting with arthralgia, Raynaud phenomenon, generalized edema, and a positive anti-small nuclear ribonucleoprotein particle antibody was diagnosed as mixed connective tissue disease. The kidney biopsy showed thrombotic microangiopathy. Bone marrow smear showed bone marrow hyperplasia and biopsy revealed suspected light chain restricted expression, megakaryocyte proliferation, and moderate to severe bone marrow fibrosis. A lymph node biopsy was conducted and the histopathological findings were consistent with the subtype of mixed Castleman disease. The clinical symptoms were relieved after regular chemotherapy. After above examination results and clinical manifestations, the final diagnoses was TAFRO syndrome. The she was started on chemotherapy with bortezomib, cyclophosphamide, and dexamethasone. After chemotherapy, symptoms such as thrombocytopenia, hematuria and proteinuria disappeared, lymphadenopathy and VEGF level decreased, and bone marrow fibrosis relieved. Our case illustrated the first cases of shared characteristics of mixed connective tissue disease and iMCD-TAFRO syndrome. Cytokines may play a role in the shared pathogenicity of the iMCD-TAFRO syndrome and systemic autoimmune diseases. Therapy directly against inflammatory factors such as corticosteroids or chemotherapy have an important therapeutic implication.

Bone Health Impairment in Patients with Hemoglobinopathies: From Biological Bases to New Possible Therapeutic Strategies.

Hemoglobinopathies are monogenic disorders affecting hemoglobin synthesis. Thalassemia and sickle cell disease (SCD) are considered the two major hemoglobinopathies. Thalassemia is a genetic disorder and one of the major hemoglobinopathies determined by an impairment of globin chain production, which causes an alteration of erythropoiesis, an improvement in hemolysis, and an alteration of iron homoeostasis. In SCD, the mutations are on the β-globin chain of hemoglobin which results in a substitution of glutamic acid by valine with consequent formation of Hemoglobin S (HbS). Several factors are involved in bone metabolism alteration in patients with hemoglobinopathies, among them hormonal deficiency, bone marrow hyperplasia, iron overload, inflammation, and increased bone turnover. Bone metabolism is the result of balance maintenance between bone deposition and bone resorption, by osteoblasts (OBs) and osteoclasts (OCs). An impairment of this balance is responsible for the onset of bone diseases, such as osteoporosis (OP). Therefore, here we will discuss the alteration of bone metabolism in patients with hemoglobinopathies and the possible therapeutic strategies to contain and/or counteract bone health impairment in these patients, taking into consideration not only the pharmacological treatments already used in the clinical armamentarium, but also the new possible therapeutic strategies.

Analysis of influence of MR signs on Harris score in ARCO stages 2-4 femoral head necrosis

To analysis and determine MR signs of Harris score ARCO stages 2-4 in osteonecrosis of femoral head (ONFH). Thirty-four patients with ONFH of ARCO stages 2 to 4 who underwent routine MR, T2 mapping, 3D-SPACE sequence examination and Harris score were retrospectively collected from January 2019 to June 2020, and 3 patients were excluded, and 31 patients were finally included, including 23 males and 8 females, aged from 18 to 62 years old with an average of(40.0±10.8) years old. Among them 21 patients with bilateral femoral head necrosis, totally 52 cases, including 17 with ARCO stage 2 patients, 24 ARCO stage 3, and 11 ARCO stage 4. MR imaging signs (femoral head collapse depth, ONFH index, bone marrow edema, hyperplasia, grade and T2 value of cartilage injury, and joint effusion) were scored and measured on the picture archiving and communication system (PACS) workstation, and the cartilage quantitative parameter T2 value was calculated and measured on Siemens postprocessing workstation. Pearson correlation analysis was used to evaluate the correlation between various MR signs and Harris score, and then multiple linear regression analysis was used to examine impact of MR signs on Harris hip score. Femoral head collapse depth(r=-0.563, P=0.000), grade of cartilage injury(r=-0.500, P=0.000), and joint effusion (r=-0.535, P=0.000) were negatively correlated with Harris score by Pearson correlation analysis. Multiple linear regression analysis showed that joint effusion(β=-6.198, P=0.001) and femoral head collapse depth(β=-4.085, P=0.014) had a significant negative impact on Harris hip score. Femoral head collapse depth and joint effusion both had significant negative relationship with Harris hip score. It is recommended to routinely evaluate femoral head collapse depth and joint effusion quantitatively and gradedly, so as to efficiently and accurately assist clinical diagnosis and treatment.

Lateral facial profile of β-thalassemia major in Javanese children: a photogrammetric analysis

BACKGROUND Impairment of globin chain synthesis in patients with β-thalassemia major causes ineffective erythropoiesis. This condition triggers bone marrow hyperplasia and can lead to craniofacial bone abnormalities. This study aimed to evaluate the lateral facial profile of β-thalassemia major, identify any differences when compared with a control group of similar age and ethnicity, and reveal the facial profile of β-thalassemia major in Javanese children.
 METHODS This cross-sectional study included 35 β-thalassemia major children aged 7–15 years. They were divided into 3 groups based on their age. Lateral facial photogrammetry was taken by measuring the forehead protrusion and nasolabial angle. Data were then mapped to the normal group of children in the same age group and descriptively analyzed using SPSS software.
 RESULTS The 1 and 2 SD group had a higher prevalence of β-thalassemia major in children aged 7–9 years for both sexes. The older age group had a closer mean value to those of children without thalassemia.
 CONCLUSIONS The forehead and maxillary profile of Javanese β-thalassemia major children tended to be protrusive, especially in the 7–9 years age group, while the older age groups had closer mean values to those of children without thalassemia.

Efficacy and Safety of Recombinant Thrombopoietin Combined with Immunosuppressive Therapy for Primary Treatment of Severe Aplastic Anemia: A Single-Center, Open-Label, Randomized, Clinical Trial

Standard immunosuppressive therapy (IST) combined with eltrombopag is the first-line treatment regimen for severe aplastic anemia patients who are ineligible for allo-hematopoietic stem cell transplantation (allo-HSCT). However, there was no second-generation TPO receptor agonist available in China before 2019, and only a first-generation TPO preparation was available for clinical treatment. Although rhTPO and eltrombopag act on different receptor binding sites, the differentiation pathways are similar, so rhTPO can theoretically also stimulate hematopoietic stem cells and promote hematopoietic recovery in AA. In 2015, we used rhTPO combined with IST as the first-line treatment for SAA. The results showed that the OR and quality of response in the rhTPO group were significantly higher than those in the control group. Multivariate analysis showed that the application of rhTPO was an independent prognostic factor affecting the efficacy of IST. Based on these results, we performed a single-center, randomized, open-label, parallel-controlled clinical trial to assess the hematologic response rate and long-term safety 3, 6, and 12 months after treatment. The study was conducted between 2015 and 2019 with 188 patients with primary SAA who were randomly assigned to the control or rhTPO group (95 with IST alone and 93 with rhTPO). For rhTPO, 15000 IU was administered subcutaneously starting on day 14 after p-ATG administration and then once every other day for a total of 14 injections. The median age of the 93 patients in the rhTPO group was 33 years (range 16-67). These included 58 patients with SAA and 35 patients with very severe aplastic anemia (VSAA), which were comparable to the control group. There were no significant differences in baseline clinical parameters between groups. No significant differences were observed in the overall hematologic response rates at 3, 6, and 12 months between the rhTPO and control groups (3 months: 40.9% vs 41.1%, P = 0.979; 6 months: 52.7% vs 52.6%, P = 0.994; 12 months: 65.6% vs 63.2%, P = 0.782, respectively). The median time of platelet transfusion independent was 49 (0-376) days in the rh-TPO group and 47 (0-195) days in the control group, which was not different between the two groups ( P = 0.757). The median time of RBC transfusion independence was 47 (5-404) days and 49 (0-295) days, respectively ( P = 0.592). The median follow-up time was 64 (1.3-86.2) months. The 5-year overall survival was 86.4% (95% confidence interval [CI], 77.2-92.0%) in the rh-TPO group and 84.1% (95% CI, 74.5-90.3%, P = 0.898) in the control group. The 5-year event-free survival was 51.7% (95% CI, 41.0-61.5%) and 54.1% (95% CI, 43.5-63.6%, P = 0.947) in the two groups, respectively. No significant differences were observed in the relapse rate and clonal evolution rate between the two groups (11.5% vs. 11.7%, 4.3% vs.3.2%, repectively). Subcutaneous rhTPO injection was well tolerated by patients, with no significant adverse effects other than mild localized pain at the injection site. The patients in the rhTPO group did not have concomitant thrombotic or embolic complications; moreover, reticulo-fibrillary or collagen fibrillary hyperplasia in bone marrow was not detected in any patients at three and six months after IST. Ten patients had anti-TPO antibodies at the end of TPO treatment, with no positive anti-TPO antibodies. In conclusion, our study is the only prospective randomized controlled clinical trial in China of rhTPO combined with IST for the treatment of SAA, with no difference in the hematologic response rate of the 28-day rhTPO combination regimen compared to standard IST; however, we did confirm the safety of rhTPO at this dose. Further clinical studies are required to determine the most effective dosage and duration of rhTPO treatment.

Impaired Immunosuppressive Effect of Bone Marrow Mesenchymal Stem Cell-Derived Exosomes on T Cells in Aplastic Anemia

Background: Previous studies have verified the dysfunction of mesenchymal stem cells (MSCs) for immunoregulation in acquired aplastic anemia (AA) patients. Exosomes derived from MSCs can partially substitute MSC acting as immune regulator. Dysfunction of exosomes (Exos) derived from AA-MSC ( AA-Exos) may play a key role in immunologic dissonance. Objectives: This study aims to investigate the interaction between exosomes derived from mesenchymal stem cells and T cells of patients with aplastic anemia by examining the immunosuppressive effects of exosomes on T cells both in vitro and in vivo. Besides, this study intends to explore the differently expressed miRNA of exosomes and mRNAs of T cells between patients with AA and healthy donors (HD) to better understand the mechanisms of AA on molecular lever. Method: In this study, CD3 + T cells were collected and cocultured with AA-Exos and exosomes derived from HD-MSC (HD-Exos). The proliferation, differentiation and activation of CD3 + T cells were detected to compare the immunosuppressive effects between AA-Exos and HD-Exos. An immune-mediated murine model of AA was structured to compare the therapeutic effect of AA-Exos and HD-Exos. Furthermore, total RNA including miRNA from exosomes we purified and total RNA of CD3 + T cells were extracted for RNA-seq in order to construct the miRNA-mRNA network for interactions and functional analysis. Results: Compared to HD-Exos, AA-Exos exhibit a weakened ability to inhibit the proliferation of CD3+/CD4+/CD8+ T cells (40.73% vs 32.43%，p = 0.0007；46.20% vs 38.08%，p = 0.0009；36.28% vs 29.73%，p = 0.0077). Furthermore, AA-Exos show reduced capacity to suppress T cell activation compared to HD-Exos. In AA-Exos group, the ability of inhibiting CD4+ T cell differentiation towards Th1 is diminished (12.80 vs 10.17%， p = 0.0047), while the induction of Th17 cell generation is significantly enhanced (3.613% vs 1.153%， p=0.0015). Additionally, AA-Exos demonstrate weaker capabilities to inhibit CD8+ T cell differentiation towards Tc1 compared to HD-Exos (41.93% vs 35.40%，p=0.0007). The capacity of AA-Exos to induce Treg is also weakene (7.78% vs 10.36%，p=0.041).HD-Exos other than AA-Exos can rescued the AA mice. Administration of HD-Exos effectively improves the bone marrow failure, the BMNC counting revealed that HD-Exos had more potency in ameliorating bone marrow hyperplasia than AA-Exos (8.27×106/L vs 0.85×106/L，p=0.0021). The RNA cargo of exosomes was believed to be a key mediator exerting their function, especially the carry of miRNA.Importantly, we identified some differentially expressed miRNA involved in immune response under the standard of adjusted p < 0.05 &| log2 (foldchange) | > 1.5, such as miR-199, miR-128, miR-486 and miR-375. The Gene Ontology analysis of differentially expressed genes (DEGs) revealed involvement of various cellular processes, such as lymphocyte chemotaxis, lymphocyte migration and response to interferon-gamma. The Kyoto Encyclopedia of Genes and Genomes analysis illustrated upregulation of critical pathways associated with T cell function after co-culturing with AA-Exos compared with HD-Exos, such as graft-versus-host disease, Th17 cell differentiation, and JAK-STAT signaling pathway. A miRNA-mRNA network was established to visualize the interaction between them, in which a total of 61 node and 142 edge network pathways were constructed, including 14 miRNAs and 47 mRNAs. To validate the findings, qPCR was performed on four DEGs, and significant differences were observed in three of them, namely MRPL40, TNF and CCL4L2 (p=0.0116, p=0.0016, p=0.0122, respectively). Conclusion: In summary, AA-Exos had impaired immunosuppressive effect on T cells, less ability to rescue AA mice and differently expressed miRNA profile, which might partly account for the pathogenesis of AA as well as provide a new target of AA treatment.

Allogeneic Hematopoietic Stem Cell Transplantation for Classical Paroxysmal Nocturnal Hemoglobinuria

Classical paroxysmal nocturnal hemoglobinuria (cPNH) is a rare acquired clonal hematopoietic cell disease. Eculizumab is effective in PNH patients, though, it must be administered indefinitely. As the technology of allogeneic hematopoietic stem cell transplantation (allo-HSCT) improves in recent years, what is the position of HSCT in treatment of cPNH? This study discusses the efficacy and safety of allo-HSCT in cPNH. The clinical data of 5 cPNH patients who underwent allo-HSCT from 2019 to 2022 were collected. All patients were diagnosed cPNH according to guidelines from International PNH Interest Group. Indications for HSCT were transfusion-dependent and nonavailability of eculizumab. The cohort was all male, with a median age of 26 years (range, 26 to 46 years) at the time of transplantation. All patients had documented hemolysis and no thrombosis during monitoring. The median time from diagnosis to allo-HSCT was 5.5 years (range, 3.6 to 18 years). The median PNH granulocyte clone size was 96.3% (range, 90% to 99.7%), and the median lactate dehydrogenase (LDH) level was 2224IU/L (8.896-fold of the upper limit of normal). All patients were detected bone marrow hyperplasia by trephine biopsy. The stem cell source came from 4 haploidentical donors and 1 HLA-identical sibling donor. Among the 5 patients, 3 patients underwent myeloablative conditioning (MAC) and 2 patients underwent reduced-intensity conditioning (RIC) treatment. None of the patients experienced primary implantation failure. Neutrophil implantation time was 15 (13-21) days, and platelet implantation time was 24 (13-60) days. 3 patients developed grade II acute graft-versus-host disease (aGVHD). No patients developed grade Ⅲ-Ⅳ aGVHD. 2 patients developed localized chronic graft-versus-host disease (cGVHD) involving lung and skin respectively, and no patients developed extensive cGVHD. PNH clones turned negative in all patients after 2 months (range, 1 to 3 months) of transplantation. At a median follow-up of 16 (6-34) months, 1 patient died of relapse and infection. The remaining 4 patients survived, of which 2 patients had discontinued all drugs. cPNH has its own unique clinical features. Allo-HSCT could cure cPNH patients who can withdrawal from treatment of all medication. Haploidentical donor HSCT is an acceptable choice for cPNH, and RIC constitutes a valid option.

Importance of FDG-PET (Fluorodeoxyglucose Positron Emission Tomography) in Staging and Response Assessment of Hairy Cell Leukemia (HCL)

Alessandro Mancini and Luca De Carolis are Co-first authors. Nicodemo Baffa, Brunangelo Falini and Enrico Tiacci are Co-last authors. INTRODUCTION HCL is an indolent BRAF-mutated leukemia usually presenting with cytopenias, splenomegaly, little or no lymphadenopathy, heavy bone marrow (BM) infiltration and few circulating leukemic cells. Imaging exams for staging include abdominal echography and computed tomography (CT), while the role of PET is largely unknown. METHODS 84 patients (pts) with relapsed/refractory HCL participating to our clinical trials of BRAF inhibitor-based therapies prospectively underwent PET/CT pre- and post-treatment, in addition to usual examinations for response assessment. Since FDG uptake is normally higher in liver than spleen, BM and lymph nodes, uptake in the latter tissues was considered abnormal if equal to or higher than the liver. Splenomegaly was defined by a longest spleen diameter >13 cm on PET/CT. The standard definition of complete remission (CR) in HCL just requires no palpable splenomegaly on physical examination, in addition to resolution of cytopenias as well as no hairy cells visible in the blood smear and the BM biopsy by non-immunological stains. RESULTS Pre-therapy, 58/77 (75%) non-splenectomized pts showed abnormal splenic FDG uptake (maximal standardized uptake value/SUVmax: median 4.8, vs 3.9 for the liver), which was always diffuse without focal lesions. Conversely, focal splenic uptake is not infrequent in splenic marginal zone lymphoma (18%, 7/39 pts - Abdom Radiol 2018;43:2721; vs of 0/58 HCL pts, p=0.0003), which may aid in the differential diagnosis with this HCL-mimicker. Moreover, 14/58 (24%) HCL pts. with a PET+ spleen did not have splenomegaly, and splenic uptake returned to normal in 10/11 evaluable cases (91%) achieving at least a partial response post-therapy, which suggests leukemic spleen involvement even without splenomegaly. Abnormal diffuse BM uptake was observed in 41/82 evaluable pts (50%; median SUVmax 5, vs 3.4 for the liver). In these cases, leukemic involvement in the BM biopsy was greater (median 80%) than in the other 41 pts (median 65%; p-value <0.01), pointing to HCL infiltration as the cause of abnormal BM uptake rather than to reactive hyperplasia of non-involved BM. Post-therapy, pathologic spleen uptake normalized in 46/51 evaluable pts (90%), and their CR rate was higher (32/46 pts, 70%) than in the other 5 pts remaining PET+ (1/5, 20%; p=0.047). Among the 46 PET- cases, 31 (67%) were not splenomegalic while 15 (33%) had residual splenomegaly (up to 17 cm; Fig. 1A); interestingly, within PET- cases relapse-free survival (RFS) was similar not only in conventionally defined CR cases with (n=8) vs without (n=24) residual non-palpable splenomegaly (p=0.84 after a median follow-up of 19 months), but also when the 6 cases meeting all conventional CR criteria except for still palpable splenomegaly were added to the 8 CR pts with residual non-palpable splenomegaly (p=0.7 after a median follow-up of 20.5 months - Fig. 1B). These findings suggest that metabolic status reflects splenic HCL involvement more reliably than spleen size or palpability, a concept that could improve the current definition of CR in HCL. Abnormal BM uptake resolved after treatment in 34/37 evaluable pts (92%), and their CR rate was again higher (23/34, 68%) than in the other 3 pts remaining PET+ (0/3; p=0.047). Leukemic infiltration of the BM biopsy was lower in the 34 PET- vs the 3 PET+ pts (median 5% vs 70%-80%, respectively; p=0.0004), indicating that BM abnormal uptake reflects HCL infiltration load rather than reactive BM hyperplasia post-therapy. Finally, abnormal lymph node uptake was observed pre-therapy in 9/84 (11%) pts (median SUVmax 7.1, vs 3.4 for the liver) and normalized after treatment in 7/8 evaluable pts (88%), including one case with residual lymph node enlargement (2.2 cm); all these 7 pts had achieved a CR while the remaining pt had no response to therapy. CONCLUSIONS PET is potentially useful in the differential diagnosis with splenic marginal zone lymphoma and may aid in the clinical management of HCL pts by detecting metabolic involvement of the main disease sites even in the absence of organomegaly. Importantly, PET signal tracks with response to therapy even in case of persistent organomegaly, which may lead to refine the definition of CR in HCL.

Multiplex allele-specific polymerase chain reaction-based prevalence study of canine trapped neutrophil syndrome in Thailand.

Trapped neutrophil syndrome (TNS) is an autosomal recessive genetic disorder found in Border Collies and is characterized by peripheral neutropenia and myeloid hyperplasia of bone marrow. The underlying cause of TNS is associated with a 4-base pair deletion mutation in the vacuolar protein sorting 13 homolog B (VPS13B) gene. In this study, we proposed and validated a novel multiplex allele specific-polymerase chain reaction (MAS-PCR) technique to assess the prevalence of TNS using VPS13B genotypes of Border Collies and Thai Ridgebacks in Thailand. We assessed the prevalence of TNS in 100 Border Collies and 30 Thai Ridgebacks using MAS-PCR-based allelic discrimination technique of the VPS13B gene. We then confirmed the VPS13B genotypes by direct DNA sequencing. A total of 130 samples were successfully genotyped using MAS-PCR assays. Of the two dog breeds examined, the VPS13B mutation was present in Border Collies, whereas Thai Ridgebacks were unaffected by this mutation. In Border Collies, 96% of dogs tested had an intact VPS13B genotype, whereas the remaining individuals had a heterozygous mutation genotype, with prevalence and mutated VPS13B allele frequencies of 4% and 2%, respectively. Using a novel MAS-PCR assay targeting the VPS13B gene, this study demonstrates for the first time that carriers of TNS exist in Border Collies in Thailand. This assay is a reliable and cost-effective tool for diagnosing TNS based on VPS13B genotypes and is suitable for routine clinical practice.

Состояние костномозгового кроветворения у больных хронической лучевой болезнью, облученных с различной мощностью дозы

Purpose: to study the features of the state of the bone marrow according to cytological and histological studies at different periods of the course of CRD, caused by exposure to different dose rates, which developed as a result of professional prolonged exposure in a cohort of former employees of a radiation hazardous enterprise who underwent inpatient examination at the clinic of the A.I. Burnazyan Federal Medical and Biomedical Center in the period up to 1995. Material and methods: the study of the results of cytological and histological studies of the bone marrow was carried out in former workers of the Mayak Production Association who were exposed to long-term industrial exposure with a dose rate of 0.008−0.07 Gy / day (15 people), 0.003−0.007 Gy / day (12 people) and less than 0.001 Gy/day (25 people), during periods of formation, outcomes, immediate and long-term consequences of CRS. At the stage of studying the results of histological examination of the bone marrow, 54 more patients with CRS were added to the third group, irradiated with a dose rate of less than 0.001 Gy / day. Statistical processing of the material was carried out using the IBM SPSS Statistics software package.23 using the Kruskal‒Wallis test and the Mann‒Whitney U-test for independent samples. The results obtained were considered statistically significant at p < 0.05. Results: At a dose rate of 0.008−0.07 Gy / day during the formation period, the myelogram revealed narrowing of the granulocyte and expansion of the red germs, acceleration of maturation of granulocytes with a normal erythrocyte maturation index and leuko-erythroblast ratio. In peripheral blood – agranulocytosis. In the period of outcomes and immediate consequences − narrowing of the granulocytic, expansion of erythrocyte germs, acceleration of maturation of neutrophils with other myelogram parameters within normal limits. In the blood − agranulocytosis, anemic syndrome. In the long term, in the case of restoration of hematopoietic function, narrowing of the granulocytic germ with other myelogram parameters within the reference values. However, with a similar course of CRS in 60 % of patients in periods of outcomes and long-term consequences, the development of myelodysplastic syndrome with transformation into acute leukemia or aplastic anemia is possible. At an irradiation dose rate of 0.003−0.007 Gy / day in the period of CRD formation, the myelogram revealed an expansion of the granulocytic germ, with other indicators within the normal range. In the period of outcomes and immediate consequences, an acceleration of maturation of granulocytes and an increase in the leuko-erythroblastic ratio were found. In the long term, a narrowing of the granulocytic and expansion of the erythrocyte sprouts, a slight acceleration of the maturation of granulocytes were found. Hystological examination: polymorphocellular bone marrow – in 11 out of 25 patients, hypoplasia – in 9 out of 25, signs of hyperplasia – in 5 out of 25. In the long term, 2 patients from this group developed oncohematological diseases. At an irradiation power of less than 0.001 Gy / day, during all periods of CRD, normal values of granulocytic and erythrocyte sprouts were noted in the myelograms of patients. In the period of CRD formation, the normal size of the granulocytic germ was achieved due to the accelerated maturation of neutrophils. The leuko-erythroblastic ratio in the period of long-term consequences was significantly higher than the norm (5.29 and 4.5, respectively). Hystological examination: 32 out of 64 patients had polymorphocellular bone marrow, 22 out of 64 had hypoplastic bone marrow, 7 out of 64 had bone marrow hyperplasia. Conclusion: regular changes in hematopoietic tissue and peripheral blood in CRD can serve as diagnostic criteria, based on which it is possible to assume the dose rate of radiation to which the patient was exposed, and also on their basis, it is possible to predict the outcome and long-term consequences of CRD that developed as a result of this exposure.

Evaluation of Mandibular Bone Microstructure in CT Scans of People with Sickle Cell Disease.

Sickle cell disease (SCD) is a common inherited disease, and is characterized by a genetic modification that determines the production of a hemoglobin with altered morphology. This anatomical change of hemoglobin leads to vaso-occlusive disorders and premature hemolysis of the cell, causing chronic anemia and bone marrow hyperplasia due to increased hematopoietic demand. As a consequence, several skeletal changes are reported in the skull, spine, ribs, pelvis, femur, and metatarsals. In the craniofacial region, dentofacial deformities are described, especially maxillary protrusion. However, studies evaluating bone microarchitecture are scarce. The aim of this study is to evaluate the mandibular bone microstructure of people with SCD on computed tomography (CT) scans. Morphometric parameters were analyzed on CT scans of the mandible of people with SCD and people without this disease or any other disease affecting bone metabolism, matched for sex and age. The results were compared by Student's t-test for paired samples and for an error probability of 5%. This study demonstrated that the mandibular bone of people with SCD presents significantly less number, connectivity and thickness of bone trabeculae, as well as having a lower fractal dimension and greater porosity. Mandibular bone of people with SCD has lower bone density and more widely spaced trabeculae.

Cytopenia: a report of haplo-cord transplantation in twin brothers caused by a novel germline GATA1 mutation and family survey.

Cytopenia due to the abnormal regulation of GATA1 could manifest as varying degrees of thrombocytopenia and/or anemia and more severely in male children than in female children. Here, we describe the case of pancytopenic and transfusion-dependent twin brothers at our center whose bone marrow puncture revealed low bone marrow hyperplasia. Whole-exome sequencing revealed that the twins had a new germline GATA1 mutation (nm_002049: exon 3:c.515 T >C:p.F172S), which confirmed the diagnosis of GATA1 mutation-related pancytopenia. The mutation was inherited from their mother, who was heterozygous for the mutation. Sanger sequencing verified the pathogenicity of the mutation. Further family morbidity survey confirmed that GATA1 mutation-related pancytopenia is an X-linked recessive genetic disorder. We developed haploid hematopoietic stem cell transplantation programs for twins, with the father as the only donor, and finally, the hematopoietic reconstruction was successful. Although they experienced acute graft-versus-host disease, hemorrhagic cystitis, and a viral infection in the early stage, no abnormal manifestations or transplant-related complications were observed 3months after transplantation. Through hematopoietic stem cell transplantation technology for one donor and two receptors, we eventually cured the twins. The p.F172S variant in the new germline GATA1 mutation may play an essential role in the pathogenesis of GATA1 mutation-related cytopenia.

Efficacy of CLAE Chemotherapy Regimen Followed by Allogeneic Hematopoietic Stem Cell Transplantation in Patients with Relapsed/Refractory Acute Leukemia

To observe the efficacy and safety of CLAE intensive chemotherapy followed by allogeneic hematopoietic stem cell transplantation (allo-HSCT) in patients with relapsed/refractory acute leukemia (R/R AL). CLAE regimen [cladribine 5 mg/(m2·d), d 1-5; cytarabine 1.5 g/(m2·d), d 1-5; etoposide 100 mg/(m2·d), d 3-5] followed by allo-HSCT was used to treat 3 R/R AL patients. The patients received CLAE chemotherapy in relapsed or refractory status and underwent bone marrow puncture to judge myelodysplastic state. After an interval of 3 to 5 days, followed by preconditioning regimen for allo-HSCT [fludarabine 30 mg/(m2·d), d -7 to d -3; busulfan 0.8 mg/kg q6h, d -6 to d -3 or d -5 to d -2. If the bone marrow hyperplasia was not active and the blasts were less than 10%, busulfan should be used for 3 days. If the bone marrow hyperplasia was active and the blasts were more than 10%, busulfan should be used for 4 days]. Cyclosporin A, mycophenolate mofetil and short-term methotrexate were used for graft-versus-host disease (GVHD) prevention. After transplantation, the status of minimal residual disease (MRD) and bone marrow chimerism were regularly monitored in all 3 patients, and demethylation drugs or dasatinib were used to prevent recurrence 3 months after transplantation. 2 patients with t(11;19) translocation and relapse/refractory acute myeloid leukemia recurred within 6 months after induction of remission, and received intensive chemotherapy with CLAE regimen followed by haploidentical allo-HSCT and unrelated donor allo-HSCT, respectively. The two patients both relapsed 6 months after transplantation, then achieved complete remission by donor lymphocyte infusion, interferon, interleukin-2 and other methods, and disease-free survival was 2 years after transplantation. The other patient was chronic myelogenous leukemia who developed acute lymphoblastic leukemia during oral administration of tyrosine kinase inhibitor, accompanied by T315I and E255K mutations in ABL1 kinase region and additional chromosomal abnormalities. After morphological remission by induction chemotherapy, central nervous system leukemia was complicated. Intensive chemotherapy with CLAE regimen followed by sibling allo-HSCT was performed in the positive state of MRD. The patient relapsed 3 months after transplantation, and achieved remission after chimeric antigen receptor T-cell (CAR-T) therapy, however, he died 5 months after transplantation because of severe cytokine release syndrome (CRS) and GVHD. CLAE regimen followed by allo-HSCT may be an effective salvage treatment option for R/R AL patients to prolong the overall survival.

Effect of Recombinant Human Thrombopoietin (rhTPO) on Hematopoietic Reconstitution in Allogeneic Hematopoietic Stem Cell Transplantation Model

To explore the effect of recombinant human thrombopoietin (rhTPO) on hematopoietic reconstruction in allogeneic hematopoietic stem cell transplantation (allo-HSCT) model. The C57BL/6 mice were employed as the donors, and BALB/c mice as recipients. The bone marrow mononuclear cells of the donor mice were extracted and pretreated, which then were injected with 5×106 per mouse through the tail vein of the recipient to establish an allo-HSCT model. The implantation of hematopoietic stem cells in the recipient mice was detected by flow cytometry on the 28th day after transplantation. Next, the successfully modeled recipient mice were randomly divided into experimental group and control group. The rhTPO was injected into mice in the experimental group on the first day after transplantation, while the saline was injected into mice in the control group. Both groups were injected for 14 consecutive days. The peripheral blood and bone marrow hematopoiesis of the two groups were observed on day 1, 3, 7, 14, and 21 after transplantation. The expression rate of H-2Kb in the bone marrow of recipient mice was 43.85% (>20%) on the 28th day after transplantation, which indicated that the recipient mice were successfully chimerized. Meanwhile, counts of PLTs on the day 3, 7, 14, and 21 after transplantation in the experimental group were higher than those in the control group with statistical significances (P<0.05). In addition, hematopoietic function of bone marrow was suppressed in both groups on day 1, 3 and 7 after transplantation, but hematopoietic bone marrow hyperplasia was better in the experimental group than in the control group. On day 14 and 21 after transplantation, the hematopoietic function of bone marrow in the two groups was recovered, and the experimental group showed more obvious than the control group. rhTPO can effectively stimulate the production of PLTs and facilitate the recovery of white blood cells and hemoglobin after allo-HSCT, and promote hematopoietic recovery and reconstitution of bone marrow.

SARS-CoV-2 infection augments species- and age-specific predispositions in cotton rats

Heterogeneity of COVID-19 manifestations in human population is vast, for reasons unknown. Cotton rats are a clinically relevant small animal model of human respiratory viral infections. Here, we demonstrate for the first time that SARS-CoV-2 infection in cotton rats affects multiple organs and systems, targeting species- and age-specific biological processes. Infection of S. fulviventer, which developed a neutralizing antibody response and were more susceptible to SARS-CoV-2 replication in the upper respiratory tract, was accompanied by hyperplasia of lacrimal drainage-associated lymphoid tissue (LDALT), a first known report of mucosa-associated lymphoid tissue activation at the portal of SARS-CoV-2 entry. Although less permissive to viral replication, S. hispidus showed hyperplasia of bone marrow in the facial bones and increased pulmonary thrombosis in aged males. Augmentation of these features by SARS-CoV-2 infection suggests a virus-induced breach in regulatory mechanisms which could be devastating for people of all ages with underlying conditions and in particular for elderly with a multitude of ongoing disorders.

The Hematopoietic Protective Effect of PDGF-BB on Radiation-Induced Myelosuppression Model Mice

To investigate the hematopoietic protective effect of platelet-derived growth factor (PDGF)-BB on radiation-induced myelosuppression model mice and effect of anti-apoptosis of megakaryocyte line Meg-01 cells, and its possible mechanism. Mice were radiated with 4 Gy of 137Csγ ray to establish the model of myelosuppression. Mice were weighed and peripheral blood cell were counted before radiation (day 0) and day 7, 14 and 21 after radiation. On the 21 st day, the mice were killed. The sternal tissues of the mice were taken for morphological observation, and the femoral bone marrow cells were cultured for the assay of colony cell forming units (CFU). Meg-01 cells were cultured without FBS for 24 h to induce apoptosis, and then treated with PDGF-BB for 48 h. The effects of PDGF-BB on the proliferation were investigated by cell counting. Flow cytometry was used to detect early apoptosis (Annexin V), mitochondrial membrane potential (JC-1) and the expression of caspase-3. Peripheral blood cell counts of mice showed that PDGF-BB stimulated the recovery of white blood cells, red blood cells and platelets after radiation (P<0.05), especially for white blood cells. Morphological examination showed bone marrow hyperplasia in PDGF-BB group, the numbers of megakaryocytes and their progenitor cells were higher than those in the control group. PDGF-BB significantly stimulated the formation of CFU-MK, CFU-GM, BFU-E and CFU-F. PDGF-BB showed a strong proliferation effect in the concentration range of 5-50 ng/ml (P<0.001). PDGF-BB (50 ng/ml) significantly reduced the positive expression of Annexin V (P<0.01). The mitochondrial membrane potential in the control group was decreased when compared with PDGF-BB group, which indicated that the number of apoptotic cells was increased (P<0.01). Besides, the expression of caspase-3 in PDGF-BB group was significantly lower than that in control group (P<0.05). PDGF-BB has a protective effect on the hematopoietic system of myelosuppression model mice, especially megakaryocytes and their progenitor cells. PDGF-BB has pro-proliferative and anti-apoptotic effects on Meg-01 cells, and the mechanism may be mediated through JC-1 and caspase-3 pathway.

Diagnosis and genetic analysis of polycythemia in children and a novel EPAS1 gene mutation.

Unlike in adults, there is no consensus on management and diagnosis of polycythemia in children. This study aims to evaluate the diagnosis and verify the algorithm in children with polycythemia. Seventy-nine children with polycythemia were followed-up in our pediatric hematology-oncology clinic between December 15, 2019, and July 15, 2021. After eliminating secondary causes (hypoxia, pulmonary, cardiac diseases), we checked for genetic mutations, including congenital erythrocytosis gene panel (JAK, EPOR, EPAS1, EGNL1, HBB, HBA, BPGM, and VHL). We also compared parameters for secondary and idiopathic polycythemia groups. Of the 79 children, thirty-five had secondary polycythemia (hypoxia, pulmonary, cardiac diseases), and one was diagnosed with a novel likely pathogenic mutation c.2089C>G; p.Pro697Ala in exon 13 of EPAS1 gene. Others (n=35) had persistent and idiopathic polycythemia. Here, we compared the idiopathic and secondary cases. We found that the ratio of family history of polycythemia (n=4 (9.5%) vs 0%, respectively) was higher in the second group (p=0.009). In addition, the mean age (14.7±3.52 vs 13.4±4.67 respectively) (p=0.042) and the ratio of erythroid hyperplasia in bone marrow [n=3 (8.6%) vs 0% respectively] (p=0.003) was higher in the idiopathic polycythemia group, compared to secondary polycythemia patients. Finding the genetic defect in polycythemia is a significant issue. Due to being a rarity in children, the first line JAK mutation analysis should be performed in selected cases. This study is the first description of a Turkish patient with EPAS1 p.Pro697Ala mutation, thereby expanding our knowledge about the clinical features of the disease. However, new investigations are required to confirm its function.

Animal Model Establishment and Its Mechanism of Cytarabine-Iduced Myelosuppression

To establish an optimized model of bone marrow suppression induced by cytarabine (Ara-C) in C57BL/6 mice and preliminarily explore the mechanism of myelosuppression based on the cycle and apoptosis of BMNC. C57BL/6 mice were intraperitoneally injected with Ara-C 50, 100 and 200 mg/kg for 7 days, respectively. The survival rate and body weight of C57BL/6 mice were monitored. The number of peripheral blood cells and bone marrow nucleated cells (BMNC) was detected, and the morphology of bone marrow, thymus and spleen were measured on the 7th, 14th and 21st day of the experiment. The cycle and apoptosis of BMNC were also detected by flow cytometry. Ara-C 200 mg/kg caused 46.7% mortality in mice, and other doses had no significant effect on mortality. All doses of Ara-C induced bone marrow suppression in mice, as shown by a decrease in the number of peripheral blood cells (WBC, Neu, RBC, PLT) and BMNC (P<0.05), decrease in bone marrow hyperplasia, accompanied by immunosuppression and compensatory hematopoiesis of the spleen, and the above manifestations and duration were dose-dependent. Among them, the myelosuppression caused by Ara-C 50 mg/kg recovered quickly, and caused by Ara-C 200 mg/kg was too severe. The result of flow cytometry showed that Ara-C could cause S and G2/m arrest and increased apoptosis in BMNC. Ara-C can induce myelosuppression in mice with a dose-dependent severity and duration, and the model of myelosuppression with Ara-C 100 mg/kg is more optimized. The mechanism is related to the inhibition of BMNC proliferation and the promotion of apoptosis.

Dioscin Regulating Bone Marrow Apoptosis in Aplastic Anemia.

BackgroundAplastic anemia (AA), a disease of bone marrow failure, is caused by CD8+T mediated apoptosis of hematopoietic cells. However, traditional immunosuppressive therapy (IST) has severe liver and kidney toxicity and even cannot achieve the expected therapeutic effect in some patients.PurposeOur study is aimed to investigate the effect and mechanism of dioscin (DNS) for treating AA.MethodsBriefly, we established and evaluated the AA mouse model, DNS and positive control drugs were used for intervention treatment. After 14 days of intervention, femoral bone marrow pathology, bone marrow mononuclear cells (BMMCs) apoptosis rate, bone marrow CD34+ cell surface Fas (CD95) expression and Fas signaling pathway key proteins were detected.ResultsAfter the establishment of the AA mouse model, the number of peripheral blood cells including granulocytes, erythrocytes, hemoglobin, platelets and reticulocytes in the AA group model was significantly decreased compared with the group control (P < 0.01). The degree of bone marrow hyperplasia in the sternum and femur is extremely low. After different drug interventions, compared with the group model, the number of peripheral blood cells in the AA mice rebounded significantly in group DNS (P < 0.01). Not only that the apoptosis rate of BM-MCs decreased (P < 0.01), meanwhile, the CD95 molecule expressed on the CD34+ bone marrow cells had a significant decline (P < 0.01), and the expression level of the key proteins of Fas signaling pathway was also significantly decreased (P < 0.01).ConclusionDNS recovered the peripheral pancytopenia and bone marrow failure in AA mice. DNS reduced the key protein of Fas signaling pathway level to inhibit apoptosis of bone marrow cells to treat AA.