The utility of methylmalonic acid, methylcitrate acid, and homocysteine in dried blood spots for therapeutic monitoring of three inherited metabolic diseases.

Backgroud: Routine metabolic assessments for methylmalonic acidemia (MMA), propionic acidemia (PA), and homocysteinemia involve detecting metabolites in dried blood spots (DBS) and analyzing specific biomarkers in serum and urine. This study aimed to establish a liquid chromatography-tandem mass spectrometry (LC-MS/MS) method for the simultaneous detection of three specific biomarkers (methylmalonic acid, methylcitric acid, and homocysteine) in DBS, as well as to appraise the applicability of these three DBS metabolites in monitoring patients with MMA, PA, and homocysteinemia during follow-up. Methods: A total of 140 healthy controls and 228 participants were enrolled, including 205 patients with MMA, 17 patients with PA, and 6 patients with homocysteinemia. Clinical data and DBS samples were collected during follow-up visits. Results: The reference ranges (25th-95th percentile) for DBS methylmalonic acid, methylcitric acid, and homocysteine were estimated as 0.04-1.02 µmol/L, 0.02-0.27 µmol/L and 1.05-8.22 µmol/L, respectively. Following treatment, some patients achieved normal metabolite concentrations, but the majority still exhibited characteristic biochemical patterns. The concentrations of methylmalonic acid, methylcitric acid, and homocysteine in DBS showed positive correlations with urine methylmalonic acid (r = 0.849, p < 0.001), urine methylcitric acid (r = 0.693, p < 0.001), and serum homocysteine (r = 0.721, p < 0.001) concentrations, respectively. Additionally, higher levels of DBS methylmalonic acid and methylcitric acid may be associated with increased cumulative complication scores. Conclusion: The LC-MS/MS method established in this study reliably detects methylmalonic acid, methylcitric acid, and homocysteine in DBS. These three DBS metabolites can be valuable for monitoring patients with MMA, PA, and homocysteinemia during follow-up. Further investigation is required to determine the significance of these DBS biomarkers in assessing disease burden over time.

Regulation of soluble epoxide hydrolase in renal-associated diseases: insights from potential mechanisms to clinical researches.

In recent years, numerous experimental studies have underscored the pivotal role of soluble epoxide hydrolase (sEH) in renal diseases, demonstrating the reno-protective effects of sEH inhibitors. The nexus between sEH and renal-associated diseases has garnered escalating attention. This review endeavors to elucidate the potential molecular mechanisms of sEH in renal diseases and emphasize the critical role of sEH inhibitors as a prospective treatment modality. Initially, we expound upon the correlation between sEH and Epoxyeicosatrienoic acids (EETs) and also addressing the impact of sEH on other epoxy fatty acids, delineate prevalent EPHX2 single nucleotide polymorphisms (SNPs) associated with renal diseases, and delve into sEH-mediated potential mechanisms, encompassing oxidative stress, inflammation, ER stress, and autophagy. Subsequently, we delineate clinical research pertaining to sEH inhibition or co-inhibition of sEH with other inhibitors for the regulation of renal-associated diseases, covering conditions such as acute kidney injury, chronic kidney diseases, diabetic nephropathy, and hypertension-induced renal injury. Our objective is to validate the potential role of sEH inhibitors in the treatment of renal injuries. We contend that a comprehensive comprehension of the salient attributes of sEH, coupled with insights from clinical experiments, provides invaluable guidance for clinicians and presents promising therapeutic avenues for patients suffering from renal diseases.

Development of a Novel Multiplex PCR Method for the Rapid Detection of SARS-CoV-2, Influenza A Virus, and Influenza B Virus.

Objective: A sensitive and specific multiplex fluorescence rapid detection method was established for simultaneous detection of SARS-CoV-2, influenza A virus, and influenza B virus in a self-made device within 30 min, with a minimum detection limit of 200 copies/mL. Methods: Based on the genome sequences of SARS-CoV-2, influenza A virus (FluA), and influenza B virus (FluB) with reference to the Chinese Center for Disease Control and Prevention and related literature, specific primers were designed, and a multiplex fluorescent PCR system was established. The simultaneous and rapid detection of SARS-CoV-2, FluA, and FluB was achieved by optimizing the concentrations of Taq DNA polymerase as well as primers, probes, and Mg2+. The minimum detection limits of the nucleic acid rapid detection system for SARS-CoV-2, FluA, and FluB were evaluated. Results: By optimizing the amplification system, the N enzyme with the best amplification performance was selected, and the optimal concentration of Mg2+ in the multiamplification system was 3 mmol/L; the final concentrations of SARS-CoV-2 NP probe and primer were 0.15 µmol/L and 0.2 µmol/L, respectively; the final concentrations of SARS-CoV-2 ORF probe and primer were both 0.15 µmol/L; the final concentrations of FluA probe and primer were 0.2 µmol/L and 0.3 µmol/L, respectively; the final concentrations of FluB probe and primer were 0.15 µmol/L and 0.25 µmol/L, respectively. Conclusion: A multiplex real-time quantitative fluorescence RT-PCR system for three respiratory viruses of SARS-CoV-2, FluA, and FluB was established with a high amplification efficiency and sensitivity reaching 200 copies/mL for all samples. Combined with the automated microfluidic nucleic acid detection system, the system can achieve rapid detection in 30 minutes.

Normal triglyceride concentration and the risk of diabetes mellitus type 2 in the general population of China.

Introduction: Hypertriglyceridemia and its derivatives are independent predictors of diabetes mellitus type 2 (T2DM). However, the relationship between triglyceride concentrations within the normal range and the incidence of T2DM remains to be clarified. This study investigated the potential relationship between variations in plasma triglyceride levels within the normal range and T2DM onset using data from a longitudinal study of health and retirement in China. Methods: Between, 2010 and, 2016, we conducted a retrospective cohort study involving 36,441 individuals with normal triglyceride levels. Using a Cox proportional hazards regression model, we examined the connection between normal triglyceride levels and T2DM incidence. We employed this method with smooth curve fitting to investigate potential nonlinear associations. Subgroup analyses were performed based on age, sex, body mass index, smoking and drinking status, hypertension, and family history of diabetes. Results: A significant linear relationship was observed between normal triglyceride levels and the incidence of T2DM. The hazard ratio for T2DM in individuals with normal triglycerides was 1.81 (95% confidence interval: 1.39, 2.36); P<0.001). Kaplan-Meier analysis further demonstrated a prospective association between the higher tertiles of normal triglyceride levels and the development of T2DM (P<0.001). Subgroup analysis revealed a stronger positive correlation between normal triglyceride levels in females and the risk of T2DM. Discussion: An increase in triglyceride levels within the normal range is related to a continuous increase in the incidence of T2DM in the general population. These findings show that almost everyone can benefit from reducing triglyceride levels, further emphasizing the importance of lifestyle changes in the general population.

Serum circPRDM5 as a novel diagnostic biomarker for acute myocardial infarction.

BACKGROUND: Circular RNA (CircRNA) is known to play an important role in cardiovascular diseases, but its use as a biomarker of acute myocardial infarction (AMI) has not been studied. This study explores the feasibility of circPRDM5 as a novel biomarker of AMI. METHODS: CircPRDM5 was screened by bioinformatics, the correct circPRDM5 primers were tested by agarose gel electrophoresis (AGE) and Sanger sequencing, and the expression level of serum circPRDM5 was detected by Quantitative Reverse Transcription-Polymerase Chain Reaction. (qRT-PCR), and the diagnostic value of circPRDM5 was analyzed by the receiver operating characteristic (ROC) curve. RESULTS: The expression of circPRDM5 in serum of AMI patients was significantly decreased compared with that of healthy control group and angina group (P < 0.001). The area under ROC curve of serum circPRDM5 was 0.862 [95 % CI, 0.814-0.909]. The combined diagnosis of serum circPRDM5, cardiac troponin T (cTnT) and creatine kinase-MB (CK-MB) could improve the sensitivity of diagnosing AMI. The expression level of serum circPRDM5 increased after percutaneous coronary intervention (PCI). CONCLUSIONS: CircPRDM5 can be used as a novel biomarker for AMI, and its combination with cTnT and CK-MB can improve diagnostic value.

Tangshen formula improves diabetic nephropathy in STZ-induced diabetes rats fed with hyper-methionine by regulating the methylation status of kidney.

BACKGROUND: The objective of this study was to examine and analyze differential methylation profiles in order to investigate the influence of hyper-methioninemia (HM) on the development of diabetic nephropathy (DN). Male Wistar rats, aged eight weeks and weighing 250-300 g, were randomly assigned into four groups: a control group (Healthy, n = 8), streptozocin-induced rats (STZ group, n = 8), HM + STZ group (n = 8), and the Tangshen Formula (TSF) treatment group (TSF group, n = 8). Blood glucose levels and other metabolic indicators were monitored before treatment and at four-week intervals until 12 weeks. Total DNA was extracted from the aforementioned groups, and DNA methylation landscapes were analyzed via reduced representative bisulfite sequencing. RESULTS: Both the STZ group and HM + STZ group exhibited increased blood glucose levels and urinary albumin/creatinine ratios in comparison with the control group. Notably, the HM + STZ group exhibited a markedly elevated urinary albumin/creatinine ratio (411.90 ± 88.86 mg/g) compared to the STZ group (238.41 ± 62.52 mg/g). TSF-treated rats demonstrated substantial reductions in both blood glucose levels and urinary albumin/creatinine ratios in comparison with the HM + STZ group. In-depth analysis of DNA methylation profiles revealed 797 genes with potential therapeutic effects related to TSF, among which approximately 2.3% had been previously reported as homologous genes. CONCLUSION: While HM exacerbates DN through altered methylation patterns at specific CpG sites, TSF holds promise as a viable treatment for DN by restoring abnormal methylation levels. The identification of specific genes provides valuable insights into the underlying mechanisms of DN pathogenesis and offers potential therapeutic targets for further investigation.

CircRNA and ferroptosis in human disease: Insights for new treatments.

Circular RNA (circRNA), classified as a type of non-coding RNA, has gained significant attention in the field of biology due to its distinctive ring structure and functional properties. Recent research has provided evidence that specific circRNAs have the ability to modulate disease progression through diverse mechanisms, one of which is by regulating cellular ferroptosis. Ferroptosis is a form of regulated cell death that is driven by iron dependency and lipid peroxidation, and extensive investigations have revealed a relationship between ferroptosis and disease development. In addition to evidence that both circRNAs and ferroptosis exert critical roles in disease progression, circRNAs have also been shown to actively mediate the process of ferroptosis. The relationship between circRNAs and ferroptosis therefore influences disease progression and offers novel targets for disease treatment. By directly or indirectly modulating the expression of circRNAs that regulate the expression of ferroptosis-related proteins, it may be possible to impact disease progression by promoting or inhibiting ferroptosis. Current research indicates such approaches may hold significant value in a wide variety of common diseases across physiological systems. This review comprehensively summarizes the findings of recent studies investigating the roles of circRNAs in the regulation of ferroptosis in various diseases.

Role of HLA class I and II alleles in susceptibility to ankylosing spondylitis in Chinese Han.

OBJECTIVE: The objective of the study was to clarify the associations of HLA class I and II alleles with ankylosing spondylitis (AS) among Chinese Han. METHODS: We performed HLA genotyping and Sanger sequencing for 68 HLA-B*27(-), 62 HLA-B*27(+) AS patients, and 70 controls. Case-control analyses and separate analyses of HLA-B*27(-) patients were performed. One-way ANOVA and Kruskal-Wallis multiple comparisons test were used to analyze the effects of HLA-A\B\C\DRB1\DQB1 alleles on clinical characteristics of HLA-B*27(-) and HLA-B*27(+) patients. RESULTS: In the HLA-B*27(+) group, positive associations were seen with A*11:02, B*27:04, B*27:05, C*02:02, C*12:02, and DRB1*04:01 and negative associations were seen with A*33:03, B*07:02, B*57:01, and C*07:02. The age at onset was greater in HLA-B*27(-) patients than in HLA-B*27(+) patients (30.03 ± 15.15 vs. 23.08 ± 7.79 years). In the HLA-B*27(-) group, those with A*01:01, B*13:01, B*13:02, C*01:02, C*04:01, DQB1*02:01, DQB1*06:01, and DRB1*03:01 had an earlier onset than those without these alleles, while patients carrying B*40:02, C*07:02, C*12:02, C*15:02, DQB1*05:02, and DQB1*05:03 had a delayed onset. In the HLA-B*27(-) group, A*32:01(+), C*08:01(+), and DRB1*04:05(-) women were likely to develop AS. In the HLA-B*27(+) group, DQB1*03:02(+) women may be more likely to develop AS. DRB1*12:02 and HLA-B*27 interacted with the distribution of AS-affected sites. In the HLA-B*27(+) group, DRB1*12:02(+) patients were likely to have peripheral joint involvement. CONCLUSION: HLA class I and II alleles other than HLA-B*27 contribute to AS predisposition and characteristics among Chinese Han patients.

Discrimination of SARS-CoV-2 omicron variant and its lineages by rapid detection of immune-escape mutations in spike protein RBD using asymmetric PCR-based melting curve analysis.

BACKGROUND: The SARS-CoV-2 Omicron strain has multiple immune-escape mutations in the spike protein receptor-binding domain (RBD). Rapid detection of these mutations to identify Omicron and its lineages is essential for guiding public health strategies and patient treatments. We developed a two-tube, four-color assay employing asymmetric polymerase chain reaction (PCR)-based melting curve analysis to detect Omicron mutations and discriminate the BA.1, BA.2, BA.4/5, and BA.2.75 lineages. METHODS: The presented technique involves combinatory analysis of the detection of six fluorescent probes targeting the immune-escape mutations L452R, N460K, E484A, F486V, Q493R, Q498R, and Y505H within one amplicon in the spike RBD and probes targeting the ORF1ab and N genes. After protocol optimization, the analytical performance of the technique was evaluated using plasmid templates. Sensitivity was assessed based on the limit of detection (LOD), and reliability was assessed by calculating the intra- and inter-run precision of melting temperatures (Tms). Specificity was assessed using pseudotyped lentivirus of common human respiratory pathogens and human genomic DNA. The assay was used to analyze 40 SARS-CoV-2-positive clinical samples (including 36 BA.2 and 4 BA.4/5 samples) and pseudotyped lentiviruses of wild-type and BA.1 viral RNA control materials, as well as 20 SARS-CoV-2-negative clinical samples, and its accuracy was evaluated by comparing the results with those of sequencing. RESULTS: All genotypes were sensitively identified using the developed method with a LOD of 39.1 copies per reaction. The intra- and inter-run coefficients of variation for the Tms were ≤ 0.69% and ≤ 0.84%, with standard deviations ≤ 0.38 °C and ≤ 0.41 °C, respectively. Validation of the assay using known SARS-CoV-2-positive samples demonstrated its ability to correctly identify the targeted mutations and preliminarily characterize the Omicron lineages. CONCLUSION: The developed assay can provide accurate, reliable, rapid, simple and low-cost detection of the immune-escape mutations located in the spike RBD to detect the Omicron variant and discriminate its lineages, and its use can be easily generalized in clinical laboratories with a fluorescent PCR platform.

Experimental Characteristics of Patients with Newly Diagnosed Multiple Myeloma.

Background: Multiple myeloma (MM), a malignant plasma cell proliferative disease, makes up to 1% of all cancers and somewhat exceeds 10% of all hematological cancers. Since it affects many organs, the signs and symptoms of myeloma vary greatly. This investigation was carried out to identify the clinical and laboratory characteristics of MM. Method: From January 1, 2014, to June 30, 2020, 169 in-patients who received a MM diagnosis for the first time at China-Japan Friendship Hospital in Beijing had their medical information examined. Results: Among 169 newly diagnosed patients, the median age was 60 years (26-84 years). Seven patients were younger than 40 years, and 16.0% (27/169) were 70 years or older. 40.8% (69/169) had IgG M-protein and 27.2% (46/169) had IgA. 84% (142/169) of patients were in the Durie Salmon stage 3. The major sign and symptoms at diagnosis were fatigue (100/169, 59.2%), bone pain (96/169, 56.8%), and weight loss (34/169, 20.1%). Anemia was present initially in 94.0% (159/169), high erythrocyte sedimentation rate in 92.7% (101/109), and thrombocytopenia in 26.6% (45/169). Similarly, hypercalcemia, renal insufficiency, and hypoalbuminemia were observed in 19.3% (31/161), 27.8%, and 75.7% respectively. Immunoparesis was found in 94% (110/117) of IgG, IgA, or IgM patients, and in 87% (33/38) of light chain myeloma patients. A localized band was found in 78.3% (123/157) of patients upon serum protein electrophoresis while monoclonal protein was detected by immunofixation in 91.5% (139/152) of patients. 4.1% (7/169) of the patients had non-secretory myeloma. The prevalence of light chain myeloma was 22.5% (38/169), and these individuals were more likely than other myeloma patients to have renal insufficiency (50% versus 21%, P < .05). In 84.8% of patients, the bone marrow had 10% or more plasma cells. Conclusion: The notable features that can be concluded from this study are the early onset of myeloma in the Chinese population and an advanced disease stage at the time of diagnosis with most of them accompanying anemia, hypoalbuminemia, and immunoparesis.

Severe autoimmune hemolytic anemia complicating hereditary spherocytosis treated successfully with glucocorticoids and cyclosporine: a case report.

BACKGROUND: Severe autoimmune hemolytic anemia complicating hereditary spherocytosis is life threatening and has not been described in a case report. Here, we report a case in which this intractable disease was treated successfully with glucocorticoids and cyclosporine. CASE PRESENTATION: A 25-year-old female patient with hereditary spherocytosis developed severe autoimmune hemolytic anemia after respiratory syncytial virus infection. Her hemoglobin level was 26 g/L and various anti-red blood cell antibodies were detected in her serum, making blood matching difficult. Glucocorticoid monotherapy was ineffective. With the addition of cyclosporine (50 mg/12 h), the patient's hemoglobin level increased significantly and the symptoms associated with anemia were greatly relieved. CONCLUSION: In patients with severe autoimmune hemolytic anemia, especially when the presence of multiple anti-red blood cell antibodies and alloantibodies interferes with blood matching, a glucocorticoid-cyclosporine regimen may be tried.

EDTA plasma in glass and polyethylene terephthalate tubes is suitable for measurement of B-type natriuretic peptide on the Mindray CL-6000i.

AIMS: To explore differences in B-type natriuretic peptide (BNP) concentration and stability and evaluate BNP accuracy in different collection tubes. METHODS: BNP concentrations in heparin/glass, EDTA/glass, and EDTA/polyethylene terephthalate (PET) tubes were measured on the Mindray CL-6000i at 0.5, 1, 2, and 4 h after collection. Differences were evaluated using Wilcoxon's paired tests and Bland-Altman plots. BNP stability and measurement accuracies were estimated using Kruskal-Wallis H tests and recovery tests. RESULTS: BNP concentrations in EDTA/glass tubes were 31.4% higher than those in heparin/glass tubes and 3.04% lower than those in EDTA/PET tubes. BNP stability significantly decreased in the heparin/glass tube. BNP remained stable in EDTA/glass and EDTA/PET tubes at room temperature for 4 h. BNP recovery rates in heparin/glass, EDTA/glass, and EDTA/PET tubes were 77.46, 86.04, and 88.23%, respectively. CONCLUSIONS: Plasma in EDTA/glass and EDTA/PET tubes is suitable for BNP measurement on the Mindray CL-6000i.

Fast and Sensitive Detection of SARS-CoV-2 Nucleic Acid Using a Rapid Detection System Free of RNA Extraction.

Objectives: To establish and evaluate the analytical and clinical performance of the Flash20 SARS-CoV-2 nucleic acid rapid detection system free of RNA extraction. Methods: The limit of detection (LoD) was determined using a negative nasopharyngeal swab matrix spiked with different concentrations of SARS-CoV-2 virus; a total of 734,337 reference sequences of viral genomes from GenBank were used for the in-silico analysis to assess the inclusivity of the assay. The specificity of the system was evaluated by testing 27 medically relevant organisms. A total of 115 clinical specimens were collected and tested on the Flash20 SARS-CoV-2 detection system and with an FDA-approved comparator test to assess the clinical performance of the system. Results: The LoD of the Flash20 SARS-CoV-2 detection system is 250 copies/mL with a positive rate ≥90% (n = 20); alignments results showed that over 99% identity of the primer and probe of the Flash20 SARS-CoV-2 nucleic acid rapid detection system to the available SARS-CoV-2 sequences; the omicron samples tested 100% positive. None of the 27 organisms showed cross-reactivity with the Flash20 SARS-CoV-2 nucleic acid rapid detection system. Among all the 215 clinical samples, the Flash20 SARS-CoV-2 nucleic acid rapid detection system exhibits a high sensitivity of 99.24% (131/132) and 100% (83/83) specificity. Conclusion: The nucleic acid rapid detection system provides sensitive and accurate detection of SARS-CoV-2 free of RNA extraction. The high sensitivity and short time to results of approximately 35 minutes may impact earlier infection control and disease management.

Total serum vitamin B12 (cobalamin) LC-MS/MS assay as an arbiter of clinically discordant immunoassay results.

OBJECTIVES: Measurement of the serum levels of vitamin B12 (VB12) is key for evaluating VB12 deficiency-dependent anemia. Immunoassay, the major method for determining VB12, tends to give false-normal results because of the presence of anti-intrinsic factor (IF-Ab) or other factors such as heterophilic antibodies et al. This study aimed to develop a liquid chromatography tandem mass spectrometry (LC-MS/MS) method that is helpful for distinguish false normal VB12 results measured by the immunoassay. METHODS: Different forms of VB12 were derivatized into CN-B12, which was collected through solid-phase extraction and analyzed via LC-MS/MS. 236 serum samples were measured both by LC-MS/MS and immunoassay, results were compared, and the IF-Ab effect was evaluated. RESULTS: The LC-MS/MS assay afforded a linear slope from 20 to 4,000 pmol/L for CN-B12. OH-VB12, methyl-VB12, and CoA-VB12 showed recovery within 89.3-109.5%. The intra-assay CV of VB12 was 2.6-4.1%, whereas the total CV was 9.3-9.8%. Passing-Bablok regression between LC-MS/MS and immunoassay results showed that the slope was 1.085 and the intercept was -15.691. The Bland-Altman plot showed that the mean difference and difference% were -34.6 pmol/L and 0.3%, respectively. Inter-rater agreement analysis showed that the linear weighted kappa value was 0.885, implying good agreement between the two methods. However, two samples were falsely elevated and one sample was falsely normal in the immunoassay compared with LC-MS/MS. The LC-MS/MS method helped in the distinction of false-normal VB12 results shown by the immunoassay. CONCLUSIONS: The VB12 LC-MS/MS method can be used as an arbiter of clinically discordant immunoassay results.

A novel assay based on DNA melting temperature for multiplexed identification of SARS-CoV-2 and influenza A/B viruses.

Introduction: The severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) and influenza viruses can cause respiratory illnesses with similar clinical symptoms, making their differential diagnoses challenging. Additionally, in critically ill SARS-CoV-2-infected patients, co-infections with other respiratory pathogens can lead to severe cytokine storm and serious complications. Therefore, a method for simultaneous detection of SARS-CoV-2 and influenza A and B viruses will be clinically beneficial. Methods: We designed an assay to detect five gene targets simultaneously via asymmetric PCR-mediated melting curve analysis in a single tube. We used specific probes that hybridize to corresponding single-stranded amplicons at low temperature and dissociate at high temperature, creating different detection peaks representing the targets. The entire reaction was conducted in a closed tube, which minimizes the risk of contamination. The limit of detection, specificity, precision, and accuracy were determined. Results: The assay exhibited a limit of detection of <20 copies/µL for SARS-CoV-2 and influenza A and <30 copies/µL for influenza B, with high reliability as demonstrated by a coefficient of variation for melting temperature of <1.16% across three virus concentrations. The performance of our developed assay and the pre-determined assay showed excellent agreement for clinical samples, with kappa coefficients ranging from 0.98 (for influenza A) to 1.00 (for SARS-CoV-2 and influenza B). No false-positive, and no cross-reactivity was observed with six common non-influenza respiratory viruses. Conclusion: The newly developed assay offers a straightforward, cost-effective and nucleic acid contamination-free approach for simultaneous detection of the SARS-CoV-2, influenza A, and influenza B viruses. The method offers high analytical sensitivity, reliability, specificity, and accuracy. Its use will streamline testing for co-infections, increase testing throughput, and improve laboratory efficacy.

Mutant RIG-I enhances cancer-related inflammation through activation of circRIG-I signaling.

RIG-I/DDX58 plays a key role in host innate immunity. However, its therapeutic potential for inflammation-related cancers remains to be explored. Here we identify frameshift germline mutations of RIG-I occurring in patients with colon cancer. Accordingly, Rig-ifs/fs mice bearing a frameshift mutant Rig-i exhibit increased susceptibility to colitis-related colon cancer as well as enhanced inflammatory response to chemical, virus or bacteria. In addition to interruption of Rig-i mRNA translation, the Rig-i mutation changes the secondary structure of Rig-i pre-mRNA and impairs its association with DHX9, consequently inducing a circular RNA generation from Rig-i transcript, thereby, designated as circRIG-I. CircRIG-I is frequently upregulated in colon cancers and its upregulation predicts poor outcome of colon cancer. Mechanistically, circRIG-I interacts with DDX3X, which in turn stimulates MAVS/TRAF5/TBK1 signaling cascade, eventually activating IRF3-mediated type I IFN transcription and aggravating inflammatory damage. Reciprocally, all-trans retinoic acid acts as a DHX9 agonist, ameliorates immunopathology through suppression of circRIG-I biogenesis. Collectively, our results provide insight into mutant RIG-I action and propose a potential strategy for the treatment of colon cancer.

Persistent increase of carbohydrate antigen 19-9 with an unknown reason: A seven-year follow-up case.

BACKGROUND: We described a patient who exhibited a gradual increase in carbohydrate antigen 19-9 (CA19-9) concentrations for 4 years at three hospitals, with no associated clinical manifestations; however, we were unable to define the cause of this increase, forcing us to consider whether it was a false-positive result. METHODS: Given the potential for interference, this study used multiple system detection, gradient dilution, Polyethylene glycol (PEG) precipitation and heterophilic antibody blocking assay to evaluate the reliability of CA19-9 concentration increase. RESULTS: Analysis of the patient sample using multiple systems indicated that CA19-9 concentrations showed an obvious increase (154.0, and 889.2 IU/ml, respectively) using the Cobas E602 and Advia Centaur XP systems, and were within the reference ranges (<10 IU/ml) on other modules. PEG precipitation on the Cobas E602 and Advia Centaur XP systems reduced the CA19-9 concentration, as did heterophilic blocking tube (HBT-6, HBT-1) blockade. CONCLUSION: CA19-9 was incorrectly identified to increase due to the presence of heterophilic antibodies. We recommend that heterophilic antibodies should be evaluated in cases with elevated CA19-9 level but no associated clinical manifestations to prevent false positives.

Association of ABCC2 polymorphism with clopidogrel response in Chinese patients undergoing percutaneous coronary intervention.

Aim: In this study, we investigated the association between ABCC2 polymorphism and clopidogrel response as well as the associated hypothetical mechanism. Methods: Chinese patients (213) with coronary artery disease (CAD) who underwent percutaneous coronary intervention (PCI) and received clopidogrel were recruited. Thereafter, their ADP-induced platelet inhibition rates (PAIR%) were determined via thromboelastometry. Further, the single-nucleotide polymorphisms (SNPs) of ABCC2 were genotyped using high-resolution melting curve (HRM)-PCR, while CYP2C19*2 and *3 polymorphisms were genotyped via real-time PCR. Results: The allele frequencies of ABCC2 rs717620 were 74.88 and 25.12% for the C and T alleles, respectively. Further, ABCC2 rs717620 TT carriers exhibited significantly higher PAIR% values (72.60 ± 27.69) than both CT (61.44 ± 23.65) and CC carriers (52.72 ± 21.99) (p = 0.047 and p = 0.001, respectively), and ABCC2 rs717620 CT carriers showed significantly higher mean PAIR% values than ABCC2 rs717620 CC carriers (p = 0.011). However, the PAIR% values corresponding to ABCC2 rs2273697 and ABCC2 rs3740066 carriers were not different. Additionally, CYP2C19*2 AA carriers presented significantly lower PAIR% values than CYP2C19*2 GA (p = 0.015) and GG (p = 0.003) carriers, and CYP2C19*3 GA carriers also presented significantly lower PAIR% values than CYP2C19*3 GG carriers (p = 0.041). In patients with CYP2C19 extensive metabolizers (EM), ABCC2 rs717620 TT carriers showed significantly higher PAIR% values (89.77 ± 9.73) than CT (76.76 ± 26.00) and CC carriers (74.09 ± 25.29) (p = 0.040 and p = 0.009, respectively). In patients with CYP2C19 poor metabolizers (PM), ABCC2 rs717620 CC carriers showed significantly lower PAIR% values (51.72 ± 25.78) than CT carriers (75.37 ± 23.57) (p = 0.043). Furthermore, after adjusting for confounding factors, ABCC2 rs717620 was identified as a strong predictor of clopidogrel hyperreactivity. Conclusion: We proposed a new target, ABCC2 rs717620, in the efflux pathway that affects individual responses to clopidogrel. The TT allele of ABCC2 rs717620 was also identified as an independent risk factor for clopidogrel hyperreactivity, and CYP2C19*2 and *3 showed association with an increased risk for clopidogrel resistance. Additionally, ABCC2 rs717620 may affect individual responses to clopidogrel via post-transcriptional regulation and interaction with CYP2C19. These findings provide new insights that may guide the accurate use of clopidogrel.

[Phenotypes and genotypes of 78 patients with propionic acidemia].

Objective: Propionic acidemia is a rare inherited metabolic disorder caused by propionyl CoA carboxylase (PCC) deficiency. This study aims to analyze the clinical characteristics and gene variations of Chinese patients with propionic acidemia, and to explore the correlation between clinical phenotypes and genotypes. Methods: Single-center, retrospective and observational study. Seventy-eight patients of propionic acidemia (46 males and 32 females) from 20 provinces and autonomous regions were admitted from January 2007 to April 2022. Their age of initial diagnosis ranged from 7 days to 15 years. The clinical manifestations, biochemical and metabolic abnormalities, genetic variations, diagnosis, treatment and outcome were studied. Chi-Square test or Mann-Whitney U test were used for statistical analysis. Results: Among 78 cases, 6 (7.7%) were identified by newborn screening; 72 (92.3%) were clinically diagnosed after onset, and the age of onset was 2 hours after birth to 15 years old; 32 cases had early-onset disease and 40 cases had late-onset disease. The initial manifestations included lethargy, hypotonia, vomiting, feeding difficulties, developmental delay, epilepsy, and coma. Among the 74 cases who accepted gene analysis, 35 (47.3%) had PCCA variants and 39 (52.7%) had PCCB variants. A total of 39 PCCA variants and 32 PCCB variants were detected, among which c.2002G>A and c.229C>T in PCCA and c.838dupC and c.1087T>C in PCCB were the most common variants in this cohort. The variants c.1228C>T and c.1283C>T in PCCB may be related to early-onset type. The variants c.838dupC, c.1127G>T and c.1316A>G in PCCB, and c.2002G>A in PCCA may be related to late-onset disease. Six patients detected by newborn screening and treated at asymptomatic stage developed normal. The clinically diagnosed 72 cases had varied complications. 10 (12.8%) cases of them died. 62 patients improved after metabolic therapy by L-carnitine and diet. Six patients received liver transplantation because of recurrent metabolic crisis. Their clinical symptoms were markedly improved. Conclusion: The clinical manifestations of propionic acidemia are complex and lack of specificity. Newborn screening and high-risk screening are keys for early treatment and better outcome. The correlation between the genotype and phenotype of propionic acidemia is unclear, but certain variants may be associated with early-onset or late-onset propionic acidemia.

Case report: First case of pemetrexed plus cisplatin-induced immune hemolytic anemia in a patient with lung adenocarcinoma.

Background: Drug-induced immune hemolytic anemia (DIIHA) is a rare but potentially life-threatening drug-related complication. There are no previous reports of pemetrexed plus cisplatin as first-line chemotherapy for non-small cell lung cancer, resulting in DIIHA. Case presentation: In this report, a patient with advanced-stage lung adenocarcinoma developed severe immune hemolytic anemia 21 days after pemetrexed plus cisplatin chemotherapy. Laboratory findings showed severe hemolysis, including a rapid decrease in hemoglobin (HGB) and an elevated level of reticulocytes (Rets), indirect bilirubin (IBIL), and lactate dehydrogenase (LDH). A workup for the possibility of DIIHA was performed, including a direct antiglobulin test (DAT), a test in the presence of the soluble drug, and a drug-treated red blood cell (RBC) test. It showed a strongly positive (3+) result for anti-C3d but not for anti-immunoglobin G (IgG) in DAT. Enzyme-treated RBCs reacted weakly with the patient's serum and pemetrexed when complement was added. In addition, the patient's serum and normal sera were reactive with cisplatin-treated RBCs. However, eluates from the patient's RBCs and diluted normal sera were non-reactive with cisplatin-coated RBCs. Untreated and enzyme-treated RBCs reacted with the patient's serum in the presence of soluble cisplatin. In vitro serological tests suggested that complement-dependent pemetrexed antibodies and cisplatin-associated non-immunologic protein adsorption (NIPA) might combine to cause immune hemolytic anemia. The patient's anemia gradually recovered when pemetrexed and cisplatin were discontinued. Conclusion: This rare case demonstrated that complement-dependent pemetrexed antibodies and cisplatin-associated NIPA might occur simultaneously in a patient with DIIHA.