Application and Research Progress of Lineage Tracing in Differentiation Mechanism of Hematopoietic Stem Cells--Review / 中国实验血液学杂志

Hematopoietic stem cells (HSCs) reside at the top of the hierarchy and have the ability to differentiate to variety of hematopoietic progenitor cells (HPCs) or mature hematopoietic cells in each system. At present, the procress of HSC and HPC differentiating to the complete hematopoietic system under physiological and stressed conditions is poorly understood. In vivo lineage tracing is a powerful technique that can mark the individual cells and identify the differentiation pathways of their daughter cells, it takes as a strong technical system to research HSC. Traditional lineage tracing studies mainly rely on imaging techniques with fluorescent dyes and nucleic acid analogs. Recently, newly cell tracing technologies have been invented, and the combination of clonal tracing and DNAsequencing technologies have provided a new perspective on cell state, cell fate, and lineage commitment at the single cell level. In this review, these new tracing methods were introduce and discuss, and their advantages over traditional methods in the study of hematopoiesis were summarized briefly.

Association of drug resistance of Mycoplasma pneumoniae with DNA load and genotypes in children with Mycoplasma pneumoniae pneumonia / 中国当代儿科杂志

<p><b>OBJECTIVE</b>To investigate the association of drug resistance of Mycoplasma pneumoniae (MP) with DNA load and genotypes in children with MP pneumonia.</p><p><b>METHODS</b>A total of 230 children who were hospitalized and diagnosed with MP pneumonia between January 2012 and December 2016 were enrolled. Throat swabs were collected from the 230 children, and a rapid drug sensitivity assay was used to determine the sensitivity of clinical isolates of MP to nine commonly used antibacterial agents. Quantitative real-time PCR was used to measure MP-DNA load in throat swabs. PCR sequencing was used to determine the genotype of 2063 locus of the MP 23S rRNA V domain.</p><p><b>RESULTS</b>Of the 230 children, 86 (37.4%) had genotype A in 2063 locus, 134 (58.3%) had genotype G, 8 (3.5%) had genotype C, and 2 (0.9%) had genotype T. Mutant strains (genotype G+C+T) had a significantly higher MP-DNA load than wild-type strains (genotype A) (P<0.05). The strains resistant to erythromycin, azithromycin, clarithromycin, and clindamycin had a significantly higher MP-DNA load than non-resistant strains (P<0.05). MP had a high drug resistance rate to macrolide antibiotics. More than 60% of the cases with resistance to macrolides were found to have A2063G mutations. MP was rarely resistant to quinolones (less than 2%).</p><p><b>CONCLUSIONS</b>Mutations in 2063 locus of the MP 23S rRNA V domain may result in the resistance of MP to macrolides and the change in DNA load and can be used as a basis for selecting drugs for MP.</p>

Effect of mobilization bone marrow stem cells on hypoxia inducible factor-1a and epidermal growth factor in rats with renal injury induced by ischemic reperfusion / 中华实用儿科临床杂志

Objective To investigate the mobilization of granulocyte-colony stimulating factor(G-CSF) with stem cell factor(SCF) in bone marrow stem cells(BMSC),and to observe the therapeutic effects of BMSC mobilization on repairing ischemia/reperfusion(I/R) renal injury induced by hypoxia inducible factor-1α (HIF-1α) and epidermal growth factor(EGF),and to investigate the mechanism for BMSC mobilization for repairing I/R renal injury.MethodsOne hundred and sixty male Sprague-Dawley rats(age:8-10 weeks) were randomly allocated into 4 groups(40 rats in each group):control group (group A),I/R group(group B),SCF + G-CSF treatment group (group C) and SCF + G-CSF control group(group D).The expressions of CD34 + and EGF were measured by immunohistochemistry technique,and the expression of EGF mRNA was measured by reverse transcriptase-polymerase chain reaction.Results 1.After I/R injury,the renal tubular epithelial cells were in a state of degeneration,necrosis,even exfoliation.As the time prolonged,the necrotic renal tubules were repaired gradually,but the repair in group C was not good compared with that in group B.The level of group C was obviously lower than that of group B on the 5th,10th,17th and 24th day(P ＜ 0.05).2.There was no significant difference in the percentage of CD34 + cells between group A and D(P ＞ 0.05).On 5 d postoperative,the level of CD34 + cells of group B and group C was conspicuous higher than group A and D (P ＜ 0.05),and that of group C was conspicuous higher than that of group B (P ＜ 0.05).They descended gradually from the 5 th day with the time prolonged.3.The expressions of HIF-1c of group B and group C had a highly positive reaction on the 5th day postoperatively and the level of HIF-1α of group B and group C decended to the normal level with the time prolonged,and at each time,the expression of HIF-1α of group C was significantly higher than that of other groups (P ＜ 0.05).4.On the 5th day postoperative,the expression of EGF of group B and group C showed a higher positive reaction on the 5th day postoperatively and the level of EGF in group B and group C descended to the normal level with the time prolonged,and at each time the expression of EGF of group C was significantly higher than that of the other groups (P ＜0.05).5.The expressions of EGF mRNA of group B and C showed higher positive reaction on the 5th day postoperatively(P ＜ 0.05).The level of EGF mRNA of group B was higher significantly than that of group A on the 17th day postoperatively(P ＜ 0.05).The level of EGF mRNA of group C was a peak and higher than that of other groups on the 10th day postoperatively(P ＜ 0.05).The level of EGF of group B and group C descended to a normal level with the time prolonged.6.There was a positive correlation between the expressions of HIF-1α and EGF (r =0.815,P ＜ 0.01).Conclusions Combination of SCF with the mobilization of G-CSF sufficiently homed BMSC to injured renal tissue was an important factor for the recovery of injured renal tubules.Combination of SCF with G-CSF may promote the expression of HIF-lα and EGF in the kidney may be one of the mechanisms of why combination of SCF with G-CSF can promote recovery from renal ischemic reperfusion injury.

The role of FDG-PET in staging of lymphoma and evaluation of therapeutic efficiency / 中华血液学杂志

<p><b>OBJECTIVE</b>To evaluate the application of 18F-fluorodeoxyglucose positron emission tomography (FDG-PET) to the staging and detecting residual masses of lymphoma.</p><p><b>METHODS</b>The clinical data of 179 patients with lymphoma were analyzed retrospectively. The results of FDG-PET, computed tomography (CT) and bone marrow biopsy (BMB) were compared for detection of lymph node/extranodal lymphoid tissue and bone marrow infiltration. Therapeutic efficiency was assessed by International Workshop Criteria (IWC) and Revised Integrated International Workshop Criteria (IWC + PET).</p><p><b>RESULTS</b>In the detection of 286 disease focuses in 98 patients before chemotherapy, the sensitivities of FDG-PET and CT were 73% and 70% (P < 0.01) in detecting nodal focus,and 87% and 45% (P < 0.01) in detecting extranodal lymphoma respectively. In detection of 104 lesions in 81 patients after chemotherapy,the sensitivities of FDG-PET and CT were 81% and 55% respectively (P < 0.01), and the specificities were 68% and 33%, respectively (P < 0.01) in detecting residual masses. According to IWC criteria, 33 patients achieved complete response/unconfirmed complete response (CR/CRu) , and 8 (24%) relapsed. Patients with PET-positive residual masses had a relapse rate of 40%, whereas only 21% of those with no such masses relapsed. Based on IWC + PET criteria, 25 patients achieved CR, with a relapse rate of 20%. Both FDG-PET and BMB produced positive results in 133/179 (74%) patients. Twenty-two patients with positive FDG-PET results were not detected by BMB. The sensitivities and specificities of FDG-PET for BM infiltration were 52% and 83%, respectively.</p><p><b>CONCLUSIONS</b>FDG-PET is a high sensitive and specific technique in staging and detecting residual masses of lymphoma.</p>

Clinical and mutational study of a Chinese infant with isovaleric acidemia / 中华儿科杂志

<p><b>OBJECTIVES</b>Isovaleric acidemia (IVA) is an autosomal recessive inborn error of leucine metabolism caused by a deficiency of the mitochondrial enzyme isovaleryl-CoA dehydrogenase (IVD) resulting in the accumulation of derivatives of isovaleryl-CoA. IVA is considered to be a severe, potentially life-threatening disorder that manifests with acute neonatal encephalopathy in approximately half of affected individuals, and recurrent episodes of vomiting, lethargy, coma and varying degrees of developmental delay in the other half of patients. This study was conducted to investigate the clinical features and IVD gene mutations of a Chinese patient with IVA.</p><p><b>METHODS</b>The clinical features, routine laboratory data, blood amino acid and acylcarnitine profiles and urinary organic acid profiles of a patient with IVA were reviewed. Whole coding exons of IVD gene were PCR-amplified for DNA sequencing. The novel mutation c.466G > C (G127A) was confirmed by RFLP with restriction endonuclease Hph I.</p><p><b>RESULTS</b>The patient was a 2 year and 7 month-old boy. At 3 days of age, he began to show severe vomiting and acidosis. He was treated with pyloromyotomy at 10 days of age. His recurrent vomiting was not ameliorated until beginning transition to a diet that included more carbohydrate from 4 months. He had 3 recurrent severe vomiting and acidosis later and showed obvious psychomotor retardation. Blood spot acylcarnitine profiles by MS-MS demonstrated an elevation of C5-carnitine with a peak concentration of 12.89 micromol/L (< 0.5 micromol/L). Organic acid analysis of urine by GC-MS revealed a relatively high level of isovaleric glycine. Mutational analysis of the patient's IVD gene revealed heteroallelic mutations of c.149G > A (R21H) and c.466G > C (G127A) which is a novel missense mutation. G127A mutation was not detected in any of 50 normal controls.</p><p><b>CONCLUSIONS</b>From the clinical course, obvious elevation of blood C5-carnitine and urine isovaleric glycine, this patient's disorder should be classified as "metabolically severe" type of IVA which suggest that c.466G > C (G127A) mutation could severely damage the function of the IVD protein. To our knowledge, this is the first characterization of IVD gene mutations in the mainland of China.</p>

Clinical significance of dynamic monitoring of cell chimerism following allogeneic hematopoietic stem cell transplantation / 中华血液学杂志

<p><b>OBJECTIVE</b>To evaluate the relationship of chimerism status of cell subsets with engraftment, occurrence of acute graft versus host disease (aGVHD), graft rejection and disease relapse after allogeneic hematopoietic stem cell transplantation (allo-HSCT).</p><p><b>METHODS</b>Chimerism status in peripheral blood (PB) and bone marrow (BM) of 65 patients received allo-HSCT were monitored at regular intervals post-transplant. Fluorescence-activated cell sorter (FACS) was used to sort CD3(+)T lymphocytes in 65 cases, CD3(-)CD56(+)CD16(+)NK cells in 52 cases, CD15(+) granulocytes in 32 cases and CD19(+)B lymphocytes in 20 cases post transplants. The chimerism status of different lineage cells was analyzed by polymerase chain reaction amplification of short tandem repeats (PCR-STR).</p><p><b>RESULTS</b>On day +7, NK-cells donor chimerism (DC 55.5%) was higher than other cell subsets. T lymphocyte was the latest one to reach complete donor chimerism (CDC) with a median on day +21. Patients whose T lymphocytes donor chimerism was more than 70% on day +7 and more than 95% on day +14 had a high risk for acute aGVHD. In all cases except those with ALL, the decreased DC of T lymphocytes were observed before molecular or hematological relapse occurred.</p><p><b>CONCLUSION</b>Serial and quantitative T cell chimerism analysis provides a reliable and rapid screening method for the early detection of engraftment, graft rejection, disease relapse and occurrence of aGVHD, therefore, is a prognostic tool to identify patients at high risk of aGVHD and disease relapse following allo-HSCT.</p>

Effect of catechin microcapsule on the repair of DNA damage in glomerular mesangial cells induced by H2O2 / 中南大学学报(医学版)

OBJECTIVE@#To explore the effect and possible mechanism of catechin microcapsulation on the repair of DNA damage in glumreular mesangial cells (GMCs) induced by H2O2.@*METHODS@#According to H2O2 concentration, the experiment GMCs were divided into 6 groups: a control group, 50 micromol/L group, 100 micromol/L group, 150 micromol/L group, 200 micromol/L group and 250 micromol/L group. Each group was sub-divided into 3 groups: 6 h group, 12 h group and 24 h group, in order to determining the optimum dose and the best time of detecting the DNA damage in GMCs. The cultured cells were divided into 8 groups as follows: the NS control group, the H2O2 group, the catechin groups (the final concentrations were 10.0, 15.0, and 20.0 mg/L respectively) and the various catechin microcapsulation groups (the final concentrations were 10.0, 15.0, and 20.0 mg/L respectively). At the end of the experiment, hydroxy radical (OH), malonydialdehyde (MDA) and total superoxide dismutase (tSOD) concentration of supernadant in GMCs were determined by biochemistry assay, the repair of DNA damage in GMCs were detected by single cell gel electrophoresis assay.@*RESULTS@#(1)At 6th h, H2O2 of 100 micromoL/L could cause the DNA damage of GMCs, and H2O2 of 150 micromol/L could result in DNA damage significantly. (2) No difference was found in the comet span of GMCs DNA in the catechin group and catechin microcapsulation group of different concentrations, while the DNA comet tail-long in the catechin microcapsulation group was shorter than that of the catechin group(all P(s)<0.05), and the fluorescence intensity of tail in the catechin microcapsulation group was lower than that of the catechin group(all P(s)<0.01). (3)When the concentration of catechin was 10.0 mg/L, no statistical significance was obtained in the concentration of dOH-, MDA and tSOD between the catechin microcapsulation group and the catechin group; while dOH- and MDA concentrations were lower, and the tSOD was higher in the catechin microcapsulation group than that in the catechin group when the concentration of catechin was 15.0 mg/L and 20.0 mg/L(all P(s)<0.05).@*CONCLUSION@#Catechin microcapsulation can enhance the GMCs ability of repairing DNA damage,which may be due to elevating the capacity of its anti-oxidation by catechin microcapsulation.

Role of caspase-1 and cytokines activated by caspase-1 in brain injury of the developing rats following recurrent seizures / 中国当代儿科杂志

<p><b>OBJECTIVE</b>The expressions of caspase-1 and cytokines activated by caspase-1 are associated with the pathophysiology of many diseases for its proinflammatory and proapototic peculiarity. However its relationship to brain injury of developing rats following recurrent seizures has not yet been identified. This study aimed to investigate the role of caspase-1 and cytokines activated by caspase-1 in brain injury of developing rats following recurrent seizures.</p><p><b>METHODS</b>A total of 96 postnatal 20 day Sprague-Dawley rats were randomly assigned into Control and Seizure groups. Seizures were induced in the Seizure group by flurothyl inhalation daily for six days. Brain tissues were sampled at 6 hrs, and at 1, 3, and 7 days after last seizure. The expressions of caspase-1, interleukin (IL)-18 and IL-1beta mRNA in the cerebral cortex were detected by RT-PCR. The water content of the brain and the pathological changes of cortex nerve cells were observed. Brain injury was evaluated using a semiquantitative neuropathological scoring system.</p><p><b>RESULTS</b>The levels of caspase-1 and IL-18 mRNA in the cerebral cortex of the Seizure group were obviously higher than those in the Control group at 6 hrs, and at 1, 3, and 7 days after seizure (P < 0.05 or P < 0.01). The expression of IL-1beta mRNA in the Seizure group exhibited a biphasic pattern: increased significantly at 6 hrs, and at 1 and 7 days post-seizure (P < 0.01), but was not significantly different from the Control group at 3 days post-seizure. Edema, degeneration and necrosis of nerve cells in cerebral cortex, accompanying by inflammatory cell infiltration and apoptosis of nerve cells, were observed under a light microscope in the Seizure group after recurrent seizures. The water content of the brain in the Seizure group increased significantly compared with that in the Control group at 6 hrs, and at 1 and 3 days after recurrent seizures (P < 0.01). The Seizure group had significantly higher neuropathological scores than the Control group at each time point (P < 0.01).</p><p><b>CONCLUSIONS</b>Caspase-1 and cytokines activated by caspase-1 play an important role in the developing brain injury after recurrent seizures.</p>

Pathologic features and prognosis of 21 children with isolated proteinuria / 中南大学学报(医学版)

OBJECTIVE@#To discuss the pathologic features, treatment and prognosis of the children with isolated proteinuria (IP).@*METHODS@#Twenty-one children with IP were enrolled according to their renal biopsy and were followed up for 0.5 to 10 years.@*RESULTS@#Renal biopsy was performed in all children. Among them 13 were mesangial proliferation glomerulonephritis (MsPGN) (including 3 minor, 6 moderate, and 4 severe ones), 2 minimal change nephritis (MCN), 3 IgA nephropathy (IgAN) (1 in Grade I and 2 in Grade II), 2 focal segmemtal glomerulosclerosis (FSGS) and 1 endocapillary proliferative glomerulonephritis (EnPGN). Interstitial changes could be found in MsPGN and FSGS mostly, presenting interstitial fibrosis, infiltration of inflammatory cells, atrophy of renal tubule, and the vacuolar degeneration of epithelia. All children accepted the medical treatment except the EnPGN case. Fifteen children recovered with no relapse; proteinuria persisted in 3 severe MsPGN and FSGS cases; 2 got the impaired renal function accompanied by persistent proteinuria; and 1 had hypertension.@*CONCLUSION@#The different degrees of renal damage can be found in all IP children who have persistent proteinuria. Most patients can get good outcome after aggressive therapies. However, the prognosis of those with severe MsPGN and FSGS was not so optimistic, and some reno-protective treatments should be given to postpone the deterioration of the renal function.

Effect of glucocorticoid on glucocorticoid-resistant children with primary nephrotic syndrome / 中华儿科杂志

<p><b>OBJECTIVE</b>Glucocorticoid (GC) is the first therapeutic choice of primary nephrotic syndrome (PNS). The response to GC treatment is an important indicator for the outcome of PNS children. Children with GC-resistant PNS present with incomplete or no response to GC, and may herald the progression to end-stage renal failure. However, the detailed mechanism of GC-resistance or GC-sensitive effect in these PNS children has not been clearly elucidated. The previous study by the authors indicated that there was increased expression of GR beta in PBMCs in GC-resistant children with PNS, and the over expression of GR beta resulted in GC resistance via influencing the ability of GR alpha nuclear translocation. To elucidate the relationship between GR beta expression in renal and in PBMCs and the effect of glucocorticoid on glucocorticoid-resistance children with PNS, the expression of GR alpha and GR beta in renal tissue and in PBMCs were detected by immunohistochemistry.</p><p><b>METHODS</b>Forty children with PNS were divided into two groups, GC-resistant group(20) and GC-sensitive group(20), the expression of GR alpha and GR beta in renal intrinsic cells and in PBMCs were measured with the immunohistochemistry technique. A semiquantitative score was used to evaluate the injury degree of the glomeruli and tubulointerstitium.</p><p><b>RESULTS</b>Compared with GC-sensitive group, the glomerular pathologic scores (6.91 +/- 1.98) and renal tubular pathologic scores (7.12 +/- 1.62) in GC- resistant group were significantly different (P < 0.01, respectively). GR alpha expressions of renal tissue and PBMCs were higher in the control group (58.3 +/- 2.6, 59.1 +/- 7.2) than those in the GC-sensitive group (40.2 +/- 7.2 and 36.6 +/- 5.1, P < 0.01, respectively) and GC-resistant group (35.0 +/- 8.2 and 36.4 +/- 6.6, P < 0.01, respectively). GR beta expressions of renal tissue and PBMCs were higher in the GC-resistant group (13.8 +/- 3.0 and 12.1 +/- 4.1) and in the GC-sensitive group (6.5 +/- 1.9 and 5.9 +/- 1.0) than that in control group (2.3 +/- 0.4 and 3.2 +/- 1.1, P < 0.01, respectively). GR beta expressions in renal tissue and PBMCs were higher in the GC-resistant group than that in the GC-sensitive group (P < 0.01). Compared with control group, GR beta expressions in PBMCs and in renal tissue were lower than those in mild renal lesion group (5.4 +/- 2.8, 6.46 +/- 2.50), midmedium renal lesion group (8.7 +/- 2.4 and 11.4 +/- 3.7) and (17.1 +/- 0.4 and 18.7 +/- 0.7) in severe renal lesion group (F = 5.8, 15.6, P < 0.01, respectively). GR beta expression of PBMCs had a positive correlation with GR beta expression of renal intrinsic cells (r = 0.651, P < 0.01). GR beta expressions by PBMCs and renal intrinsic cells were positively correlated with renal pathologic scores (r = 0.579 and 0.623, P < 0.01, respectively).</p><p><b>CONCLUSION</b>GC-resistant children with PNS were related to the increased GR beta expression in PBMCs and renal intrinsic cells. There was no correlation between the GR alpha expressions in PBMCs and in renal intrinsic cells. Increased GR beta expression might decrease the effect of GC via inhibiting the activity of GR alpha.</p>