Cell-free DNA in Embryo Culture Media as Non-invasive Biomarker of the Quality of Embryo Cleavage

Background: The success of in vitro-fertilization (IVF) cycles is determined in large part by the quality of embryo cleavage, which in turn, is dependent on the quality of the embryo culture media (CM). Many factors can influence the quality of embryo CM, one of which is the levels of Cell Free Deoxyribonucleic acid (DNA). Understanding the association between Cell-free DNA levels in embryo CM and the quality of embryo cleavage could help improve the quality of IVF techniques. Methods: This prospective study was conducted with 96 spent CM from patients undergoing IVF cycle, in order to determine relationships of Cell-free DNA levels in embryo CM with embryo cleavage quality on day 3. After intracytoplasmic sperm injection (ICSI), 48 embryos were evaluated on day 3 of their development, according to their cell number. Day 2 and day 3 CM corresponding to each one of the embryos was analyzed, by quantitative PCR, for estimation of Cell-free DNA levels. Results: The results revealed a significant increase in Cell-free DNA levels on day 2 CM corresponding to 4 to 6 cell embryos compared to those corresponding to 7 to 8 cell embryos (p=0.04). As for day 3 CM, the results showed no significant difference between the Cell-Free DNA levels in CM of 7-8 and those of 4-6 cell embryos (p=0.4). Also, cell free DNA levels in embryo CM, were significantly higher on day 2 compared to day 3 for both 7-8 and 4-6 cell embryos (p=0.03; p=0.04). Conclusion: We conclude that cell-free DNA levels in CM might be associated with delayed embryo cleavage.

Levels of neutrophil extracellular traps in neonates with acute respiratory distress syndrome / 中国当代儿科杂志

OBJECTIVES@#To study the changes in cell free-DNA (cf-DNA), a marker of neutrophil extracellular traps (NETs), in neonates with acute respiratory distress syndrome (ARDS), and to evaluate its relationship with the severity and early diagnosis of ARDS.@*METHODS@#The neonates diagnosed with ARDS in the Affiliated Hospital of Jiangsu University from January 2021 to June 2022 were enrolled in the prospective study. The neonates were divided into mild, moderate, and severe ARDS groups based on the oxygen index (OI) (4≤OI<8, 8≤OI<16, and OI≥16, respectively). The control group was selected from jaundice neonates who were observed in the neonatal department of the hospital during the same period, and they had no pathological factors causing neonatal jaundice. Peripheral blood samples were collected on day 1, day 3, and day 7 after admission for the ARDS group, and on the day of admission for the control group. Serum cf-DNA levels were measured using a fluorescence enzyme-linked immunosorbent assay. Serum interleukin-6 (IL-6) and tumor necrosis factor-α (TNF-α) levels were measured using enzyme-linked immunosorbent assay. A Pearson correlation analysis was used to evaluate the correlation of serum cf-DNA levels with serum IL-6 and TNF-α levels.@*RESULTS@#A total of 50 neonates were enrolled in the ARDS group, including 15 neonates with mild ARDS, 25 with moderate ARDS, and 10 with severe ARDS. Twenty-five neonates were enrolled in the control group. Compared with the control group, the serum levels of cf-DNA, IL-6, and TNF-α in all ARDS groups were significantly increased (P<0.05). Compared with the mild ARDS group, the serum levels of cf-DNA, IL-6, and TNF-α in the moderate and severe ARDS groups were significantly increased (P<0.05), and the increase was more significant in the severe ARDS group (P<0.05). The serum levels of cf-DNA, IL-6, and TNF-α in all ARDS groups were significantly increased on day 3 after admission and significantly decreased on day 7 after admission compared with those on day 1 after admission (P<0.05). The Pearson correlation analysis showed that there was a positive correlation between serum cf-DNA levels and IL-6 levels as well as TNF-α levels in 50 neonates with ARDS (P<0.05).@*CONCLUSIONS@#There is an excessive expression of NETs in neonates with ARDS, and dynamic monitoring of serum cf-DNA levels has certain clinical value in evaluating the severity and early diagnosis of ARDS in neonates.

Development status and new challenges of heart transplantation / 器官移植

Heart transplantation is the primary therapeutic option for patients with end-stage heart failure. The shortage of donors has been the main limiting factor for the increasing quantity of heart transplantation. With persistent updating and introduction of novel technologies, the donor pool has been increasingly expanded, such as using the heart from older donors, donors infected with hepatitis C virus, donors dying from drug overdose or donation after cardiac death (DCD) donors, etc. Meantime, the proportion of recipients with advanced age, multiple organ dysfunction, mechanical circulatory support and human leukocyte antigen antibody sensitization has been significantly increased in recent years. The shortage of donors, complication of recipients' conditions, individualized management of immunosuppressive therapy and prevention and treatment of long-term cardiac allograft vasculopathy are all challenges in the field of heart transplantation. In this article, novel progresses on donor pool expansion, improving the quality of recipients, strengthening the diagnosis and treatment of rejection, and preventing cardiac allograft vasculopathy were reviewed, aiming to prolong the survival and enhance the quality of life of patients with end-stage heart failure on the waiting list or underwent heart transplantation.

Establishment and evaluation of a method for extracting exogenous short DNA fragments of Schistosoma japonicum from urine samples / 中国血吸虫病防治杂志

Objective To establish the method for extracting exogenous short DNA fragments of Schistosoma japonicum from urine samples, and to evaluate the efficiency of this method for extraction from urine samples treated with various methods. Methods The S. japonicum SjG28 gene fragment was selected as a target sequence, and the 81 bp short DNA fragment was amplified on the target sequence using PCR assay. Following characterization using sequencing, the short DNA fragment was added into the urine samples as an exogenous short DNA fragment. Primers and probes were designed with SjG28 as a target gene, to establish the real-time fluorescent quantitative PCR (qPCR) assay. The sensitivity of this qPCR assay was evaluated with exogenous short DNA fragments that were diluted at a 1:10 dilution ratio as the DNA template, and the specificity of the qPCR assay was evaluated with the genomic DNA of S. mansoni, S. haematobium, Babesia, Ancyiostoma duodenaie, Cionorchis sinensis, and Paragonimus westermani as DNA templates. Exogenous short DNA fragments were added into artificial and healthy volunteers’ urine samples, followed by pH adjustment, centrifugation and concentration, and the efficiency of extracting exogenous short DNA fragments from urine samples was compared with the QIAmp Viral RNA Mini Kit (Qiagen kit) and BIOG cfDNA easy kit (BIOG kit). Results An 81 bp small DNA fragment of S. japonicum was successfully prepared, and the lowest detection limit of the established qPCR assay was 100 copies/μL of the 81 bp small DNA fragment of S. japonicum. If the genomic DNA of S. japonicum, S. mansoni, S. haematobium, Babesia, A. duodenaie, C. sinensis, and P. westermani served as DNA templates, the qPCR assay only detected fluorescent signals with S. japonicum genomic DNA as the DNA template. If the pH values of artificial urine samples were adjusted to 5, 6, 7 and 8, the recovery rates were (49.12 ± 2.09)%, (84.52 ± 4.96)%, (89.38 ± 3.32)% and (87.82 ± 3.90)% for extracting the exogenous short DNA fragment of S. japonicum with the Qiagen kit, and were (2.30 ± 0.07)%, (8.11% ± 0.26)%, (13.35 ± 0.61)% and (20.82 ± 0.68)% with the BIOG kit, respectively (t = 38.702, 26.955, 39.042 and 29.571; all P values < 0.01). If the Qiagen kit was used for extracting the exogenous short DNA fragment from artificial urine samples, the lowest recovery rate was seen from urine samples with a pH value of 5 (all P values < 0.05), and there were no significant differences in the recovery rate from urine samples with pH values of 6, 7 and 8 (all P values > 0.05). Following centrifugation of artificial [(64.30 ± 1.00)% vs. (58.87 ± 0.26)%; t = 12.033, P < 0.05] and healthy volunteers’ urine samples [(31 165 ± 1 017) copies/μL vs. (28 471 ± 818) copies/μL; t = 23.164, P < 0.05]. In addition, concentration of artificial urine samples with the 10 kDa Centrifugal Filter and concentration of healthy volunteers’ urine samples with the 100 kDa Centrifugal Filter were both effective to increase the recovery of the Qiagen kit for extracting the exogenous short DNA fragment of S. japonicum (both P values < 0.01). Conclusions A method for extracting exogenous short DNA fragments of S. japonicum from urine samples has been successfully established, and the Qiagen kit has a high extraction efficiency. Adjustment of urine pH to 6 to 8 and concentration of healthy volunteers’ urine samples with the 100 kDa Centrifugal Filter are both effective to increase the efficiency of extracting exogenous short DNA fragments of S. japonicum.

Research progress on emerging biomarkers in kidney transplantation / 器官移植

The hosts could show complex and diverse immune responses to the allografts. Whether biomarkers can be employed to explain the complexity of graft immune responses and the degree of disease damage are of significance. Conventional biomarkers, such as estimated glomerular filtration rate and blood concenrtation of immunosuppressant, lack of sensitivity and specificity in accurately identifying between immune and non-immune renal allograft injuries. Although renal biopsy is the "gold standard" for the diagnosis of postoperative complications, it still has disadvantages, such as invasiveness and high price, etc. Emerging biomarkers have potential advantages in the non-invasive diagnosis of subclinical injury of renal allograft, prediction of treatment response and individualized adjustment of immunosuppressive regimen. In this article, emerging biomarkers including blood, urine and tissue biomarkers that have been applied and are expected to be applied in clinical practice in the field of kidney transplantation were reviewed, and the range of application and effect of these biomarkers were evaluated, aiming to promote appropriate and rational application of these promising emerging biomarkers in clinical practice.

The expression and clinical significance of neutrophil extracellular trapping nets in neonatal sepsis / 中国新生儿科杂志

Objective:To study the expression and significance of neutrophil extracellular traps (NETs) in neonatal sepsis.Methods:Prospective research were used in this study. Term infants with neonatal sepsis hospitalized for the first time in the Department of Neonatology, Children's Hospital of Soochow University from June 2020 to November 2020 were selected as the sepsis group. According to a ratio of about 1∶1, term infants with mild hyperbilirubinemia who were admitted in the same period, with gestational age difference less than 1 week from those in the sepsis group, and whose parents agreed to participate in the study were selected as the control group. On admission, clinical data as well as blood samples of the two groups were collected. Levels of NETs marker citrulline histone H3-DNA (CitH3-DNA) were detected by Enzyme-linked immunosorbent assay, and circulating cell-free DNA (cfDNA) was tested by the fluorescence microplate reader. General data, white blood cell (WBC), neutrophil count (NE), platelet (PLT), C- reactive protein (CRP), blood culture, CitH3-DNA and cfDNA were compared between the two groups. The diagnostic value of CITH3-DNA and cfDNA in neonatal septicemia was analyzed by the receiver operating characteristic (ROC) curve.Results:A total of 74 infants were included in the study, including 39 cases in the sepsis group and 35 cases in the control group. CitH3-DNA and cfDNA in the sepsis group were significantly higher than those in the control group [CitH3-DNA (optical density): 0.85±0.05 vs. 0.48±0.03, cfDNA (mg/L): 0.90±0.05 vs. 0.56±0.03] ( P<0.01). There was no significant correlation between CitH3-DNA and cfDNA. The level of CitH3-DNA had no correlation with gender, gestational age, age, birth weight, WBC, NE, PLT and CRP ( P>0.05). cfDNA was positively correlated with age and NE ( P<0.05), and negatively correlated with PLT ( P<0.05). Combined with CRP, the area under the ROC curve of CitH3-DNA+CRP, cfDNA+CRP, and CitH3-DNA+cfDNA+CRP were 0.947, 0.947 and 0.970 respectively, and the sensitivity to predict neonatal sepsis were 92.3%, 84.6% and 94.9% respectively, the specificity were 94.3%, 97.1% and 100% respectively, all higher than the predictive value of each index alone. Conclusions:The plasma NETs levels increase significantly in neonatal sepsis patients, especially CitH3-DNA with a strong specificity, and can be considered as a biomarker for early diagnosis of neonatal sepsis. NETs together with CRP, could drastically improve the predictive value of neonatal sepsis.

The role of MYCN and PHOX2B gene combined with plasma cell-free DNA detection in risk stratification and predicting the prognosis of high-risk neuroblastoma / 中华实用儿科临床杂志

Objective:To explore the clinical significance of the MYCN gene, PHOX2B gene and plasma cell-free DNA (cfDNA) in risk stratification and predicting the prognosis of high-risk neuroblastoma (NB). Methods:This was a prospective study involving 94 high-risk NB children admitted to Beijing Children′s Hospital, Capital Medical University from August 2017 to December 2018.Relative levels of MYCN and PHOX2B and cfDNA at diagnosis, and 4 and 6 cycles of chemotherapy were detected, and their differences were compared by the Chi- square test.Kaplan-Meier survival analysis was performed to explore their prognostic potential in high-risk NB. Results:Among the 94 high-risk NB children, 14 cases (14.9%) had MYCN amplification, 76 cases (80.8%) had positive expression of PHOX2B and 56 cases (59.6%) had cfDNA level higher than 100 μg/L.The proportion of high lactate dehydrogenase (LDH, ≥1 500 U/L) level in the MYCN gene amplification group (6/14 cases) was higher than that in the normal group (9/80 cases) ( P=0.009). The proportion of multi-site metastasis (54/76 cases) and high neuron specific enolase (NSE) level (NSE≥370 μg/L, 37/76 cases) in PHOX2B positive group were significantly higher than those in the negative group (5/14 cases, 2/14 cases) ( P=0.015, 0.020). The proportion of high LDH and high NSE in high cfDNA concentration (≥229.6 μg/L)group (13/37 cases, 28/37 cases) were significantly higher than those in low cfDNA concentration group (2/48 cases, 10/48 cases) (all P<0.001). With the decreased tumor burden during the treatment, the copy number of PHOX2B gene and cfDNA level were significantly lower than those at the initial diagnosis [0 (0-719.6) copies vs.1 723.5 (0-186 000.0) copies; 19.0 (1.1-225.5) μg/L vs.200.6 (8.0-5 247.4) μg/L, all P<0.001]. The 2-year event-free survival (EFS) rate of the MYCN gene amplification group was significantly lower than that of the normal group[(33.3±13.1)% vs.(58.5±7.1)%, P=0.020]. The 2-year EFS rate of PHOX2B positive group was significantly lower than that of the negative group[(47.9±7.1)% vs.(79.1±11.1)%, P=0.043]. EFS rate in high cfDNA concentration group was significantly lower than that in cfDNA low concentration group[(38.6±9.8)% vs.( 71.7±8.2)%, P=0.001]. After 6 cycles of chemotherapy, EFS rate in the PHOX2B positive group was significantly lower than that in the negative group [(16.7±14.4)% vs.( 60.6±6.6)%, P=0.014]; which was significantly lower in the Metaiodobenzylguanidine (MIBG) positive group than that of the negative group[(35.2±11.7)% vs.(65.8±7.1)%, P=0.037]. The MYCN gene and cfDNA concentration were not correlated with the prognosis of high-risk NB.Survival analysis of the combination of PHOX2B and MYCN gene ( PHOX2B+ /MIBG + , PHOX2B+ or MIBG + , PHOX2B-/MIBG -) showed a significant difference in the survival among three groups[0 vs.(53.6±1.2)% vs.(65.5±7.4)%, P=0.003]. Conclusions:The MYCN and PHOX2B gene and cfDNA concentration are of significance in risk stratification and predicting the prognosis of high-risk NB.Compared with the MYCN gene and cfDNA concentration, the PHOX2B gene is more suitable for monitoring the curative effect of chemotherapy on high-risk NB.A combined analysis of PHOX2B gene and MIBG before treatment can be more accurate in evaluating the treatment effect and residual lesions.

Research progress and application of donor-derived cell-free DNA in diagnosis and treatment of kidney transplantation / 器官移植

Early diagnosis and treatment of rejection after kidney transplantation play a critical role in alleviating allograft injury. Detection of donor-derived cell-free DNA (dd-cfDNA) could be performed based on the next-generation sequencing and other techniques. The content of DNA fragments derived from necrotic and apoptotic donor kidney tissues in circulating body fluids could be determined by concentration and absolute quantitative methods, which has application potential in monitoring allograft injury in clinical practice. Compared with traditional serum creatinine and other indicators, dd-cfDNA detection may monitor allograft injury from several weeks to months in advance, providing a "time window" for clinical treatment and delaying graft failure. Along with deepening research of dd-cfDNA in recent years, dd-cfDNA has captivated widespread attention due to its non-invasiveness, high sensitivity and real-time evaluation of therapeutic effect. In this article, current study evidence and conclusions related to multidimensional application of dd-cfDNA detection in diagnosis and treatment of kidney transplantation were reviewed, and the future research and clinical application direction of dd-cfDNA were discussed, aiming to provide reference for widespread application of dd-cfDNA detection in clinical practice in China.

Donor-derived cell-free DNA can discriminate acute rejection types after kidney transplantation / 中华肾脏病杂志

Objective:To explore the value of detecting plasma donor-derived free DNA (dd-cfDNA) fraction in distinguishing antibody mediated-rejection (ABMR) and T cell-mediated rejection (TCMR) of renal allografts.Methods:Patients with acute rejection confirmed by allograft biopsy in the First Affiliated Hospital of Medical College of Zhejiang University from December 1, 2017 to July 18, 2019 were retrospectively included. Based on pathological classification of Banff renal allograft rejection in 2017, the patients were divided into ABMR group and TCMR group, and the latter was subdivided into TCMR Ⅰ subgroup and TCMR Ⅱ subgroup. The second generation sequencing and target region capture were used to detect candidates' peripheral blood dd-cfDNA. The demographic and clinicopathological data of the two groups were compared. The receiver operating characteristic curve (ROC) was used to evaluate the differential value of plasma dd-cfDNA and serum creatinine levels in two kinds of acute renal allograft rejection.Results:A total of 60 patients with acute rejection of renal transplantation were enrolled in this study, including 42 patients in TCMR group and 18 patients in ABMR group. The plasma dd-cfDNA percentage (%) in the ABMR group was significantly higher than that in the TCMR group [2.33(1.19, 4.30)% vs 0.98(0.50, 1.82)%, P=0.001]. The absolute value of dd-cfDNA in ABMR group was obviously higher than that in TCMR group [0.94(0.60, 2.27) ng/ml vs 0.43(0.20, 0.96) ng/ml, P=0.003]. ROC analysis to discriminate TCMR from ABMR showed that, the area under the curve ( AUC) of dd-cfDNA% was 0.76(95% CI 0.64-0.88), when the threshold was 1.11%, the sensitivity and specificity were 88.89% and 59.52%, respectively; the AUC of absolute value of dd-cfDNA was 0.74(95% CI 0.61-0.86), when the threshold was 0.53 ng/ml, the sensitivity was 88.89% and the specificity was 54.76%. TCMR subgroups were further analyzed, there was no significant difference between TCMR subgroups on the absolute value and percentage of dd-cfDNA (both P>0.05); dd-cfDNA% in ABMR group was apparently higher than that in TCMRⅠ subgroups ( P=0.008) and TCMRⅡsubgroup ( P=0.030). The absolute value of dd-cfDNA in ABMR group was significantly higher than that in TCMRⅠsubgroups ( P=0.003). Conclusion:Plasma dd-cfDNA level may help to distinguish between ABMR and TCMR rejection.

The application value and research progress of urine free DNA in tumor molecular diagnosis / 中华检验医学杂志

Urine cell-free DNA (ucfDNA) contains DNA fragments released from the lysis of cells in the urinary system, and circulating cell-free DNA (ccfDNA) can also enter the urine after glomerular filtration, and the mutation information carried by tumor DNA contained in ucfDNA and ccfDNA is consistent. Therefore, as a biomarker for molecular diagnosis of urological and non-urinary tumors, ucfDNA, has become a research hotspot in the field of liquid biopsy in recent years. UcfDNA has potential application value in individualized treatment, early diagnosis, dynamic monitoring of therapeutic efficacy, and prognosis assessment, etc. However, in order to realize the clinical application of urine ucfDNA, the extraction and accurate detection of ucfDNA still need to be solved.

Prediction of bloodstream infection pathogens in hematopoietic stem cell transplantation patients by plasma microbial cell-free DNA sequencing / 中华检验医学杂志

Objective:To predict the pathogens of bloodstream infection (BSI) in hematopoietic stem cell transplantation (HSCT) patients by plasma microbial cell-free DNA (mcfDNA) sequencing with and without additional amplification.Methods:A total of 978 HSCT patients were enrolled in Peking University People′s Hospital from March to July 2021, and the 7 428 blood samples were prospectively collected from pretransplant conditioning period to 4 months after transplantation. The plasma samples were separated and then cryopreserved. According to blood culture results and whether there were plasma samples before BSI onset, twenty-eight HSCT patients with positive blood culture (39 plasma samples within 1-8 days before BSI onset) and 9 HSCT patients with negative blood culture (9 plasma samples) were filtered. The 39 samples were performed with mcfDNA additional and non-additional amplification sequencing, and the 9 samples were only performed with additional amplification sequencing. With the blood culture results as the gold standard, the consistency between the sequencing and the blood culture results was observed. Student t test and Wilcoxon test were used for statistical analysis. Results:Without additional amplification sequencing, only 7 samples sequencing results were consistent with the blood culture results, and the total pathogen detection rate was 17.95% (7/39). The rates within 3 days and 4-8 days were 23.81% (5/21) and 2/18, respectively. The main pathogenic type detected was gram-negative bacteria (5/7). With additional amplification sequencing, the total pathogen detection rate was 59.26% (16/27) and the rate within 3 days was 8/13. The number of gram-positive bacteria detected was elevated (13/16) and the number of additional microorganisms in additional amplification sequencing was increased significantly ( P=0.001 0), compared with non-additional amplification sequencing. Moreover, additional sequencing analysis of 9 samples from patients with negative culture result showed that no pathogen was detected in six samples, and the common Torque teno virus in HSCT patients was detected in only three samples. Conclusion:The pathogen detection rate of plasma mcfDNA additional amplification sequencing was better than that of non-additional amplification sequencing in HSCT patients before BSI onset, especially in the first three days, which has the potential to predict BSI pathogens.

Research advances of urinary cell-free DNA in the detection and therapy of bladder cancer / 中华泌尿外科杂志

Bladder cancer is a common malignancy in the genitourinary system and the current therapeutic approaches are unsatisfactory. Urinary cell-free DNA (ucf DNA) has the ability to give comprehensive and crucial information on cancer as it carries genetic messages from cells shedding directly into urine as well as transporting from circulation. The ucf DNA of patients with bladder cancer carries disease information, suggesting that ucf DNA may have the ability to detect, monitor, and prognosticate patients with bladder cancer. The ucf DNA analysis bridges the gap between current techniques and enhances diagnostic and detection capabilities, and has a very promising future in term of translation into clinical practice. This article reviewed the progress of clinical applications of ucf DNA in bladder cancer.

Urinary donor-derived cell-free DNA as a non-invasive biomarker for BK polyomavirus-associated nephropathy / 浙江大学学报（英文版）（B辑：生物医学和生物技术）

BK polyomavirus-associated nephropathy (BKPyVAN) is a common cause of allograft failure. However, differentiation between BKPyVAN and type I T cell-mediated rejection (TCMR) is challenging when simian virus 40 (SV40) staining is negative, because of the similarities in histopathology. This study investigated whether donor-derived cell-free DNA (ddcfDNA) can be used to differentiate BKPyVAN. Target region capture sequencing was applied to detect the ddcfDNAs of 12 recipients with stable graft function, 22 with type I TCMR, 21 with proven BKPyVAN, and 5 with possible PyVAN. We found that urinary ddcfDNA levels were upregulated in recipients with graft injury, whereas plasma ddcfDNA levels were comparable for all groups. The median urinary concentrations and fractions of ddcfDNA in proven BKPyVAN recipients were significantly higher than those in type I TCMR recipients (10.4 vs. 6.1 ng/mL,

Research highlights of renal transplantation in 2020: voice from China / 器官移植

Renal transplantation is the optimal approach to improve the quality of life and restore normal life for patients with end-stage renal diseases.With the development of medical techniques and immunosuppressants, the shortterm survival of renal graft has been significantly prolonged, whereas the long-term survival remains to be urgently solved.Renal ischemia-reperfusion injury (IRI), acute rejection, chronic renal allograft dysfunction, renal fibrosis and other factors are still the major problems affecting the survival of renal graft.Relevant researches have always been hot spots in the field of renal transplantation.Meantime, 2020 is an extraordinary year.The novel coronavirus pneumonia (COVID-19) pandemic severely affects the development of all walks of life.Researches related to renal transplantation have also sprung up.In this article, the frontier hotspots of clinical and basic studies related to renal transplantation and the COVID-19 related researches in the field of renal transplantation in China were reviewed, aiming to provide novel therapeutic ideas and strategies.

Diagnostic criteria and research progress on heart allograft pathology / 器官移植

With the improvement of surgical technique of heart transplantation and clinical application of potent immunosuppressant, the quantity of heart transplantation and the survival time of heart allograft have been significantly improved. However, a series of complications, such as right ventricular failure, ischemia-reperfusion injury, acute rejection, "Quilty lesion", infection and chronic rejection characterized by transplant coronary artery disease (TCAD) may still occur at different stages after heart transplantation. The application of endomyocardial biopsy (EMB) makes it possible to observe and understand the pathological features of multiple complications of heart allograft including rejection, which has become the most accurate diagnostic tool for postoperative complications. In this article, the brief history of heart allograft pathology, main postoperative complications and pathological diagnostic criteria, and cutting edge research progress on diagnostic criteria of rejection were illustrated, aiming to bring clinical benefits to more recipients undergoing heart transplantation.

Assessment and monitoring of immune risk of kidney transplantation rejection / 器官移植

Kidney transplantation is the most efficacious treatment for end-stage renal failure. Although the shortterm survival and functional recovery of the kidney graft have been significantly improved, the long-term survival of the kidney graft remains to be enhanced. Antibody-mediated rejection (AMR) and T cell-mediated rejection (TCMR) caused by immune factors are still the most critical causes of kidney graft failure. In this article, the immune risk assessment and monitoring of kidney transplant recipients during the awaiting period, before and after kidney transplantation were reviewed. Through the evaluation of preexisting human leukocyte antigen (HLA) antibodies and non-HLA antibodies, HLA matching, lymphocytotoxicity cross-matching and immune memory cells in the recipients before kidney transplantation, programmed biopsy of the kidney graft of the recipients after kidney transplantation and monitoring of HLA antibodies, non-HLA antibodies and donor-derived cell-free DNA (dd-cfDNA), individualized immunosuppressive treatment and monitoring regimes could be established, and the incidence of rejection could be prevented, timely detected and diagnosed. According to the immune monitoring results, ineffective treatment or over-treatment could be avoided, thereby improving the long-term survival of kidney graft.

Current status of diagnosis and treatment of chronic lung allograft dysfunction / 器官移植

Chronic lung allograft dysfunction (CLAD) is the largest obstacle to the long-term survival of lung transplant recipients, which represents a series of complicated clinical manifestations of significant and persistent deterioration of lung allograft function after surgery. Due to lack of effective strategies for early diagnosis and prevention, over half of lung transplant recipients will develop CLAD within postoperative 5 years, which is likely to increase to 75% within postoperative 10 years. At present, no drug can be administered to completely prevent or reverse the progression of CLAD. In recent years, since the definition, diagnosis and treatment of CLAD have been updated by International Society of Heart and Lung Transplantation (ISHLT) in 2019, the understanding of CLAD has been significantly deepened within the international community. In this article, comprehensive diagnostic methods and potential treatment strategies of CLAD were explicitly illustrated, aiming to provide theoretical reference and insights for early monitoring and management of the incidence and progression of CLAD.

Evaluation of circulating cell free DNA in plasma as a biomarker of different thyroid diseases / Avaliação do DNA circulante livre de células no plasma como um biomarcador de diferentes doenças da tireoide

Abstract Introduction: Many studies have been done on proteomics, genomics, epigenetic, immunogenetics in many body fluids. Among these, circulating cell-free DNA (ccfDNA) entered the literature in 1948, but it has not been studied for many years due to technological deficiencies. Following recent advances, geno-metastasis has been mentioned and new research is needed in this area. ccfDNA is known to be an important biomolecule in this regard. Objective: The presence of cell-free DNA in the circulatory system may offer a tremendous opportunity to provide novel biomarkers for thyroid diseases. This experimental study was conducted to determine the amount of ccfDNA in different thyroid diseases, then to evaluate whether the ccfDNA concentration varied between the disease groups and control group. Methods: In total, we included 121 individuals in the present study. We collected blood samples and then determined the ccfDNA concentration in plasma of collected blood samples from three groups: thyroiditis (n = 33), benign (n = 37), and malignant (n = 30) and from a control group (n = 21). Results: The median values of the ccfDNA groups were found as 1610, 1665, 1685 and 576 ng/mL for the thyroiditis, benign, malign, and control groups, respectively. Findings showed that the ccfDNA of the three groups was significantly higher than the control (p < 0.0001). Each group was compared in terms of ccfDNA and the p-values of benign-thyroiditis, benign-malign, and thyroiditis-malign were 0.09, 0.65, and 0.29, respectively. Conclusions: The clear differences between thyroid diseases and controls suggest that ccfDNA is worthy of attention as a biomarker for further evaluation of different thyroid diseases. Likewise, it might indicate a clear tendency that ccfDNA can also be used to distinguish different thyroid diseases.

Resumo Introdução: Muitos estudos foram realizados em proteômica, genômica, epigenética e imunogenética em vários fluidos corporais. Entre esses, o DNA circulante livre de células (cfDNA) despontou na literatura em 1948, mas não foi estudado por muitos anos devido a deficiências tecnológicas. Após recentes avanços, a genometástase é mencionada e novas pesquisas tornam-se necessárias nessa área. Nesse sentido, o cfDNA é conhecido por ser uma importante biomolécula. Objetivo: A presença de DNA livre de células no sistema circulatório pode oferecer uma excelente oportunidade para fornecer novos biomarcadores para doenças da tireoide. Este estudo experimental foi conduzido para determinar a quantidade de cfDNA em diferentes doenças da tireoide e então avaliar se a concentração de cfDNA variou entre os grupos com doença e o grupo controle. Método: No total, 121 indivíduos foram incluídos no estudo. Coletamos amostras de sangue e, em então, determinamos a concentração de cfDNA no plasma de amostras de sangue de três grupos: tireoidite (n = 33), benigno (n = 37) e maligno (n = 30) e de um grupo controle (n = 21). Resultados: As medianas dos valores dos grupos de cfDNA foram de 1.610, 1.665, 1.685 e 576 ng/mL para os grupos tireoidite, benigno, maligno e controle, respectivamente. Os achados mostraram que o cfDNA dos três grupos com doença era significativamente maior do que o do grupo controle (p < 0,0001). Cada grupo foi comparado em termos de cfDNA e os p-valores de benigno-tireoidite, benigno-maligno e tireoidite-maligno foram de 0,09, 0,65 e 0,29, respectivamente. Conclusões: Como resultado, as óbvias diferenças entre as doenças da tireoide e os controles sugerem que o cfDNA é digno de atenção como um biomarcador para avaliação adicional das diferentes doenças da tireoide. Da mesma forma, isso pode indicar uma clara tendência de que o cfDNA também pode ser utilizado para distinção das diferentes doenças da tireoide.

Aspectos genómicos, transcriptómicos y del diagnóstico en el síndrome de Down / Genomic, transcriptomic, and diagnostic features of Down syndrome

El síndrome de Down es causado por la presencia de una tercera copia del cromosoma 21 y fue descrito por primera vez en 1838 por Jean-Etienne-Dominique, y más tarde por John Langdon Haydon Down en 1866, mientras trabajaba como superintendente médico en el Asilo Real de Earlswood. Desde ese momento, la comunidad científica puso grandes esfuerzos en tratar de elucidar diversos aspectos que influyen en la naturaleza de esta condición, y que determinan su incidencia y factores de riesgo. De igual manera, se ha puesto interés en los genes involucrados en esta enfermedad, la relación genotipo-fenotipo, la expresión del fenotipo, la variabilidad del material genético y las consecuencias transcripcionales que se producen al tener una tercera copia, ya sea parcial o total, del cromosoma 21. Además, se han invertido esfuerzos en identificar biomarcadores y en diseñar metodologías de diagnóstico prenatal no invasivo que sean altamente eficientes para un mejor diagnóstico del síndrome de Down, y así reducir su impacto negativo en las madres gestantes, al proveerlas de información neutral y precisa acerca de vivir con un hijo con síndrome de Down, y darles autonomía en la decisión de la continuación de su embarazo

Down syndrome is caused by the presence of a third copy of chromosome 21 and was first described by Jean-Etienne-Dominique in 1838, and later by John Langdon Haydon Down in 1866, while working as a medical superintendent in the Royal Earlswood Asylum. Since, the scientific community has placed great efforts in trying to elucidate different influencing features in the nature of this condition that determine their incidence and risk factors. In addition, especial attention has been given to the genes involved in this disease, the genotype-phenotype relationship, the expression of the phenotype, the variability of the genetic material and the transcriptional consequences that are produced by having a third copy, either partial or total, of chromosome 21. Additionally, efforts have been invested in identifying biomarkers and designing noninvasive prenatal methodologies that are highly efficient for a better diagnosis of Down syndrome, in order to reduce its negative impact in pregnant mothers, by providing them with neutral and accurate information about life with a child with Down syndrome, as well as providing the autonomy in the decision to continue their pregnancy

Circulating nucleic acids in the plasma and serum as potential biomarkers in neurological disorders

Neurological diseases are responsible for approximately 6.8 million deaths every year. They affect up to 1 billion people worldwide and cause significant disability and reduced quality of life. In most neurological disorders, the diagnosis can be challenging; it frequently requires long-term investigation. Thus, the discovery of better diagnostic methods to help in the accurate and fast diagnosis of neurological disorders is crucial. Circulating nucleic acids (CNAs) are defined as any type of DNA or RNA that is present in body biofluids. They can be found within extracellular vesicles or as cell-free DNA and RNA. Currently, CNAs are being explored as potential biomarkers for diseases because they can be obtained using non-invasive methods and may reflect unique characteristics of the biological processes involved in several diseases. CNAs can be especially useful as biomarkers for conditions that involve organs or structures that are difficult to assess, such as the central nervous system. This review presents a critical assessment of the most current literature about the use of plasma and serum CNAs as biomarkers for several aspects of neurological disorders: defining a diagnosis, establishing a prognosis, and monitoring the disease progression and response to therapy. We explored the biological origin, types, and general mechanisms involved in the generation of CNAs in physiological and pathological processes, with specific attention to neurological disorders. In addition, we present some of the future applications of CNAs as non-invasive biomarkers for these diseases.