Dynamic inflammatory response in a critically ill COVID-19 patient treated with corticosteroids / 浙江大学学报·医学版

OBJECTIVE@#To investigate the effect of corticosteroids therapy on the inflammatory response in a critically ill coronavirus disease 2019 (COVID-19) patient.@*METHODS@#A 55-year old female patient with critical ill COVID-19 was admitted in Taizhou Hospital on January 19, 2020. The patient was treated with methylprednisolone 80 mg on the 2nd day after admission. Thereafter, the dose was adjusted in a timely manner and the therapy lasted for 13 days. The peripheral lymphocyte subsets (CD3T, CD4 T, CD8 T, NK cells, B cells), as well as serum levels of lymphocyte factors (IL-2, IL-4, IL-6, IL-10, TNF-α, IFN-γ) were dynamically monitored.@*RESULTS@#On D1 of admission, the numbers of peripheral blood CD3 T, CD4 T, CD8 T, and NK cells were significantly lower than the normal range. With the improvement of the disease, the numbers of CD3 T, CD8 T and CD4 T cells gradually recovered and showed a linear growth trend (linear fitting equation: =18.59+109.4, <0.05). On D2 of admission, the patient's IL-6 and IL-10 levels were significantly higher than normal values, IFN-γ was at a normal high value, and then rapidly decreased; IL-2, IL-4, and TNF-α were all in the normal range. On the D6 and D7, the IL-6 and IL-10 decreased to the normal range for the first time. On the D18, the sputum virus nucleic acid test was negative for the first time, and the fecal virus nucleic acid test was still positive; on the D20 the sputum and fecal virus nucleic acid test were both negative. On D34, the patient recovered and was discharged. At the discharge the muscle strength score of the patient was 44 and the daily life ability evaluation was 90.@*CONCLUSIONS@#In the absence of effective antiviral drugs, early use of appropriate doses of corticosteroids in critically ill patient with COVID-19 can quickly alleviate inflammatory response and improve clinical symptoms, however, it may reduce the number of T cells, and to adjust the dose in time is necessary.

Genetic study of a fetus with a de novo Xp22.33;Yp11.2 translocation / 中华医学遗传学杂志

OBJECTIVE@#To delineate cytogenetic and molecular abnormalities of a fetus carrying a de novo 46,X,der(X),t(X;Y)(p22.3;p11.2).@*METHODS@#G-banded karyotyping and next-generation sequencing (NGS) were used to analyze the fetus, his father and sister. Single nucleotide polymorphism-based arrays (SNP-array), multiple PCR and fluorescence in situ hybridization (FISH) were utilized to verify the result.@*RESULTS@#G-banded karyotyping at 320 bands showed that the fetus had a normal karyotype, while NGS has identified a 3.58 Mb microdeletion at Xp22.33 and a Y chromosomal segment of about 10 Mb at Yp11.32p11.2. With the sequencing results, high-resolution karyotyping at 550-750 bands level has determined the fetus to be 46,X,der(X)t(X;Y)(p22.3;p11.2). The result was confirmed by PCR amplification of the SRY gene, FISH and SNP-array assays. The karyotypes of his father and sister were both normal. His sister also showed no amplification of the SRY gene, and her NGS results were normal too, suggesting that the karyotype of the fetus was de novo.@*CONCLUSION@#Combined karyotyping, NGS, SNP-array, PCR and FISH assay can facilitate diagnosis of XX disorder of sex development.

Cytogenetic and molecular genetic analysis of small supernumerary marker chromosomes in fetal amniotic fluid / 中华医学遗传学杂志

<p><b>OBJECTIVE</b>To explore the origin and mechanism of small supernumerary marker chromosomes (sSMC) in order to facilitate genetic counseling.</p><p><b>METHODS</b>Chromosome karyotypes of two fetuses and their immediate family members were analyzed by conventional G banding. High-throughput whole genome sequencing was used to determine the origin of sSMCs.</p><p><b>RESULTS</b>Fetus 1 was shown to have a karyotype of 47,XY,+mar but with normal FISH and B ultrasound findings. Its father also had a 47,XY,+mar karyotype with normal FISH results and clinical phenotype. High-throughput genome sequencing revealed that fetus 1 and its father were both 46,XY,dup(21)(q11.2;q21.1) with a 6.2 Mb duplication of the long arm of chromosome 21. The fetus was born with normal phenotype and developed well. Its grandmother also had a karyotype of 46,XX,t(15;21)(q13;p13) with normal FISH result and clinical phenotype. The karyotypes of its mother and grandfather were both normal. Analysis of fetus 2 showed a 47,XY,+mar karyotype with normal FISH results. High-throughput genome sequencing suggested a molecular karyotype of 46,XX. The fetus was born with normal phenotype and developed well. The karyotypes of its parents were both normal.</p><p><b>CONCLUSION</b>Considering their variable origins, identification of sSMC should combine conventional G banding analyses with high-throughput whole genome sequencing for precise delineation of the chromosomes.</p>

Clinical significance of HLA-G in patients with gastric cancer / 中华微生物学和免疫学杂志

Objective To investigate the clinical significance of CD14+HLA-G+ monocytes in pe-ripheral blood and the soluble form of HLA-G ( sHLA-G ) in plasma among patients with gastric cancer ( GC) . Methods Blood samples were collected from 135 patients with gastric cancer ( GC group) , 150 pa-tients with chronic gastritis ( CG group) and 80 healthy controls ( HC group) . Flow cytometry analysis and ELISA were used to detect the percentages of CD14+HLA-G+ monocytes in peripheral blood samples, the concentrations of sHLA-G in plasma samples and the levels of alpha fetoprotein (AFP), cacino-embryonic antigen ( CEA) , CA19-9 and CA125 in serum samples. Mann-Whitney U test was performed to analyze the differences between different groups. The feasibility of using CD14+HLA-G+ monocytes, sHLA-G, AFP, CEA, CA19-9 and CA125 as potential biomarkers to differentiate patients with GC from those with CG or healthy subjects was assessed by using receiver operating characteristic ( ROC ) curve analysis. Results The median percentages of CD14+HLA-G+ monocytes in subjects from GC, CG and HC groups were 18. 6% (12. 1%-26. 7%), 7. 3% (4. 2%-11. 0%) and 4. 6% (3. 6%-6. 3%), respectively. The percentages of CD14+HLA-G+monocytes in the peripheral blood of patients with GC were significantly higher than those in patients with CG and healthy subjects (P<0. 001). The concentrations of sHLA-G in plasma samples collected from patients with GC [(100. 6±61. 3) U/ml) were significantly higher than those in pa-tients with CG [(59.5±19. 9) U/ml) and healthy subjects [(45. 8±23. 3) U/ml] (P<0. 001). ROC curve analysis showed that in terms of GC diagnosis, the area under ROC curve ( AUC) , cutoff value, sensi-tivity and specificity for CD14+HLA-G+monocytes and sHLA-G in plasma were 0. 893 and 0. 720, 12% and 85 U/mL, 75. 8% and 50. 5%, 86. 7% and 95. 9% (P<0. 001), respectively, which indicated that CD14+HLA-G+ monocytes and sHLA-G were better than AFP, CEA, CA19-9 and CA125 in differentiating GC from CG and HC. Moreover, the multivariate logistic regression analysis revealed that the CD14+HLA-G+ monocytes, sHLA-G in plasma as well as CA19-9 and CA125 in serum were positively correlated with the risk of GC after excluding the differences caused by age and gender factors. Conclusion The levels of CD14+HLA-G+ monocytes in peripheral blood and sHLA-G in plasma increased dramatically in patients with gastric cancer, which suggested that CD14+HLA-G+monocytes and sHLA-G might be risk factors for GC and could be used as potential biomarkers for the diagnosis of GC.

Analysis of two false positive cases from noninvasive prenatal testing / 中华医学遗传学杂志

<p><b>OBJECTIVE</b>To track and analyze two false positive cases from non-invasive prenatal testing for potential fetal aneuploidy.</p><p><b>METHODS</b>The two cases, respectively reported to have XO (+++) and T18 (1/20) XO(+), were analyzed with conventional karyotyping, fluorescence in situ hybridization (FISH) and massively parallel genomic sequencing (MPS).</p><p><b>RESULTS</b>The first fetus, who was suspected for XO(+++), was verified to have super female syndrome (47,XXX/46,XX) due to confined placental mosaicism by karyotyping of amniotic fluid cells, FISH analysis of placenta and massively parallel sequencing (MPS) of fetal tissue. The second fetus, suspected to have trisomy 18 (1/20) XO(+), was verified to have Turner syndrome by karyotyping, FISH and MPS analyses of umbilical cord blood cells. And the karyotype was 45,X[48]/46, X, der(X) del(X) (p11.21) del(X) (q13.3)[62].</p><p><b>CONCLUSION</b>Non-invasive prenatal testing carries a risk for false positive diagnosis of fetal sex chromosome and trisomy 18. Combined cytogenetic and molecular techniques are required to ensure an accurate diagnosis.</p>

Flow cytometry immunophenotyping and minimal residual disease evaluation in multiple myeloma / 中华检验医学杂志

Objective To investigate the characteristics of immunophenotyping and clinical significance of MRD analysis in MM patients. Methods Multi-parameter flow cytometry was applied to analyze the immunophenotyping of malignant plasma cells from 172 MM patients, and normal plasma cells from 16 healthy individuals. MRD was analyzed in 32 MM patients with remission. Meanwhile, the effects of MRD status on the disease relapse and patient disease free survival ( DFS ) time was evaluated by following up patients. Results The immunophenotyping of normal plasma cells were CD38, CD138, CD19 and CD45 positive, while the predominant phenotype of MM cells were CD+38( 100.0% ), CD+138( 100.0% ), CD-19 ( 167/172, 97. 1% ), CD+56( 152/172, 88.4% ) and CD-45( 166/172, 96. 5% ). The characteristic markers for MM cells were CD+38, CD-138, CD-19, CD-45 and CD+56. MRD analysis revealed that, among 32 MM patients with remission, 14 patients were MRD negative and 18 patients were MRD positive. During follow-up of 2 to 16 months, the relapse rate in MRD negative patients was significantly lower ( 4/14, 28.6% ) than that of MRD positive patients ( 13/18, 72. 2% ;χ2 =6. 03, P ＜0. 05 ). Furthermore, the DFS time was significantly longer in MRD negative patients[ 16. 23( 10. 37-21.62 )months ] than that of the MRD positive patients [ 10. 07( 3. 79-16. 20 )months,χ2 =7. 53,P ＜0. 05 ]. Conclusions CD+38, CD+138, CD-19, CD-45 and CD+56 are the characteristic markers of MM cells compared to those of the normal plasma cells. MRD analysis is a valuable prognostic factor for MM patients.

Relationship between the CD44v6 expression and ERK phosphorlation in acute myeloid leukemia patients / 中华检验医学杂志

Objective To investigate expressions of CDev6 and p-ERK1/2 in AML patients and its clinical significance.Methods Expressions of CD44v6 and p-ERK1/2 on bone marrow blasts in 152 AML patients and 22 normal controls were determined by flow cytometry.Meanwhile,the effects of CD44v6 expression(CD+44v6 and CD-44v6) on relapse rate and survival time were analyzed by following up of 129 AML patients.Results Double positive expressions of CD44v6 and p-ERK1/2 were observed in 4.5%(1/22) of the normal bone marrow blasts while single positive expression of CD44v6 and p-ERK1/2 was observed in 36.2%(55/162) and 64.5%(97/162) of AML patients,respectively.The MFI of p-ERK1/2 expression on blast cells in AML patients was [14(7 -58)],which was higher than that in normal controls [8(6 - 10),U =4.2,P＜0.01].Furthermore,MFI of p-ERK1/2 expression on blast cells in CD+44v6 AML patients was [28(15-61)] ,which was significantly higher than that in CD-44v6 AML patients [18(6- 37),U =6.7,P＜0.01].A strong correlation was obtained between CD44v6 and p-ERK1/2 expression(rs = 0.7,P＜ 0.01).Among 129 patients followed up for 4 to 65 months,the data also revealed that the relapse rate of CD-44v6 AML patients was 32.1%(26/81) ,which was much lower than that in CD44v6 AML patients [81.3%(39/48),x2 = 29.13,P＜ 0.01).And the overall survival in CD44v6 AML patients was(28.5 ± 1.8)months,which was significantly worse than that of CD44v6 AML patients [(51.2±2.0) months,x2 =48.2,P＜ 0.01).Conclusion CD44v6 expression was associated with a poor survival in AML patients,and CD44v6 might promote the expansion of leukemic blast cells by up-regulating p-ERK1/2.

Relationship between CD158j expression and phosphorylated ERK levels in CD4 + CD28null T cells in cerebral infarction patients with carotid atherosclerosis / 中华微生物学和免疫学杂志

Objective To investigate the relationship between CD158j expression and phosphorylated ERK (p-ERK) in CD4+ CD28null T cells in cerebral infarction (CI) patients- with carotid atherosclerosis and its effects on carotid atherosclerotic plaque stability. Methods Percentage of peripheral CD4+ CD28null and the expression of CD158j and perform on CD4+ CD28null cells was analyzed with flow cytometry in 106 CI patients with carotid atherosclerosis, 33 CI patients with normal carotid arteries and in 50 normal controls, respectively; p-ERK expression was assayed with flow cytometry in 36 CI patients with unstable plaque, and serum IFN-γ was detected with ELISA. The intima-media thickness (IMT) of bilateral carotid arteries in all subjects was confirmed by the colour Doppler ultrasonograph imagingResults Percentage of the CD4+ CD28null T cells, expression of CD158j and perform on CD4+ CD28null T cells and the serum IFN-γ levels was dramatically higher in CI patients than that in normal controls, respectively (all P <0.01), which was decreased in an order of CI patients with patients with unstable plaque, stable plaque, carotid artery IMT and with normal carotid artery. A strong positive correlation was observed between the CD158j expression and degree of p-ERK in CI patients with unstable plaque (P < 0. 01). Conclusion CD4+ CD28null T cells were significantly increased in CI patients with carotid atherosclerosis. CD158j might up-regulate p-ERK expression and induce the proliferation of the CD4+ CD28nullT cells; consequently, higher cytokine production such as IFN-γ produced by CD4+ CD28null T cells may cause the formation of unstable plaque.