MicroRNA miR-181d-5p regulates the MAPK signaling pathway by targeting mitogen-activated protein kinase 8 (MAPK8) to improve lupus nephritis.

BACKGROUND: Lupus nephritis (LN) is a common immune disease. The microRNA (miR)-181d-5p is a potential target for treating kidney injury. However, the therapeutic role of miR-181d-5p in LN has not been investigated. This study aimed to investigate the role of miR-181d-5p in targeting mitogen-activated protein kinase 8 (MAPK8) and stimulating the MAPK signaling pathway in LN. METHODS: RT-qPCR was performed to identify the variations in miR-181d-5p expression in peripheral blood mononuclear cells (PBMCs) obtained from 42 LN patients, 30 healthy individuals, 6 MRL/lpr mice and 6 C57BL/6 mice. Western blot was used to detect the effect of miR-181d-5p on the MAPK signaling pathway in THP-1 cells and MRL/lpr mice. Enzyme-linked immunosorbent assay (ELISA) was utilized to detect the effect of miR-181d-5p on antinuclear antibodies and inflammatory factors. A dual-luciferase reporter assay was used to verify whether miR-181d-5p directly targets MAPK8. Flow cytometry was performed to evaluate apoptosis rates in transfected THP-1 cells. RESULTS: miR-181d-5p expression was downregulated in PBMCs of LN patients (P < 0.01) and MRL/lpr mice (P < 0.05). A dual luciferase reporter assay demonstrated that miR-181d-5p inhibits MAPK8 (P < 0.01). Overexpression of miR-181d-5p inhibited the phosphorylation of p38 (P < 0.001) and p44/42 (P < 0.01). Moreover, miR-181d-5p decreased the apoptosis rate of THP-1 cells (P < 0.001), and reduced the secretion of IL-6 (P < 0.01) and TNF-α (P < 0.01). Furthermore, overexpression of miR-181d-5p decreased anti-dsDNA antibody (P < 0.05), anti-Sm antibody (P < 0.01), and fibrosis levels in MRL/lpr mice. CONCLUSION: Upregulation of miR-181d-5p showed anti-inflammatory and anti-apoptotic effects on THP-1 cells in vitro and kidney injury in vivo. These effects were achieved by miR-181d-5p targeting MAPK8 to inhibit phosphorylation of p38 and p44/42. These results may offer new insights for improving therapeutic strategies against lupus nephritis.

Rapid and Ultrasensitive Detection of Plasmodium spp. Parasites via the RPA-CRISPR/Cas12a Platform.

Microscopic examination of thick and thin blood smears stained with Giemsa dye is considered the primary diagnostic tool for the confirmation and management of suspected clinical malaria. However, detecting gametocytes is relatively insensitive, particularly in asymptomatic individuals with low-density Plasmodium infections. To complement existing diagnostic methods, a rapid and ultrasensitive point-of-care testing (POCT) platform for malaria detection is urgently needed and necessary. A platform based on recombinase polymerase amplification (RPA) followed by CRISPR/Cas12a (referred to as RPA-CRISPR/Cas12a) was developed and optimized for the determination of Plasmodium spp. parasites, particularly Plasmodium falciparum, using a fluorescence-based assay (FBDA), lateral flow test strips (LFTS), or naked eye observation (NEO). Then, the established platform was assessed with clinical malaria isolates. Under optimal conditions, the detection threshold was 1 copy/µL for the plasmid, and the limit of detection was 3.11-7.27 parasites/µL for dried blood spots. There was no cross-reactivity against blood-borne pathogens. For the accuracies of RPA-CRISPR/Cas12a, Plasmodium spp. and P. falciparum testing were 98.68 and 94.74%, respectively. The method was consistent with nested PCR results and superior to the qPCR results. RPA-CRISPR/Cas12a is a rapid, ultrasensitive, and reliable platform for malaria diagnosis. The platform requires no or minimal instrumentation for nucleic acid amplification reactions and can be read with the naked eye. Compared with similar diagnostic methods, this platform improves the reaction speed while reducing detection requirements. Therefore, this platform has the potential to become a true POCT for malaria parasites.

Prevalence and molecular characterization of common thalassemia among people of reproductive age in the border area of Guangxi-Yunnan-Guizhou province in Southwestern China.

Objectives: Thalassemia, the most common global monogenetic disorder, is highly prevalent in southern China. Epidemiological and molecular characterization of thalassemia is important for designing appropriate prevention strategies in high-risk areas, especially the border area of Guangxi-Yunnan-Guizhou province in southwestern China.Methods: We recruited 38812 reproductive age couples and screened them for thalassemia. Routine blood tests as well as hemoglobin components and levels were evaluated. In addition, suspected thalassemia were identified by gap polymerase chain reaction (Gap-PCR) and PCR-based reverse dot blot (PCR-RDB).Results: The overall prevalence of thalassemia was 26.76%. Specifically, incidences of α-thalassemia, ß-thalassemia, and concurrent α- and ß-thalassemia were 17.52%, 6.92%, and 2.32%, respectively. The diagnosed α-thalassemia anomalies were associated with six gene mutations and 25 genotypes. The ß-thalassemia anomalies were associated with 12 gene mutations and 15 genotypes. Moreover, among the 1799 concurrent mutated α- and ß-thalassemia genes, 95 different genotypes were identified. Couples in which both partners were positive for α-thalassemia and ß-thalassemia isotypes were 8.80% and 2.08%, respectively. The proportion of couples at a risk of having children with thalassemia major or intermedia was high.Conclusions: This study elucidates on the prevalence and molecular characterization of thalassemia in the border area of Guangxi-Yunnan-Guizhou provinces. These findings provide valuable baseline data for genetic counseling and prenatal diagnosis, with the overarching goal of preventing and controlling severe thalassemia.

Association between genetic variants of microRNA-21 and microRNA-155 and systemic lupus erythematosus: A case-control study from a Chinese population.

BACKGROUND: Systemic lupus erythematosus (SLE) is a common autoimmune disease, and its pathogenesis remains unclear. The alteration of genetic materials is believed to play a role in SLE development. This study evaluated the association between the genetic variants of microRNA-21 (miR-21) and microRNA-155 (miR-155) and SLE. METHODS: The SNaPshot genotyping method was used to detect the genotypes of selected SNPs in patients and controls. The expression of miR-21 and miR-155 was analyzed using reverse transcription-quantitative polymerase chain reaction (RT-qPCR). The functional annotation and the biological effects of SNPs were assessed by HaploReg V4.1 and Regulome DB V2.0 software. The Hardy-Weinberg equilibrium test was used to gather statistics, and odds ratios (ORs) and 95% confidence intervals (CIs) were evaluated by logistic regression. RESULTS: The distribution difference of TA genotype in rs767649 was observed (TA vs. T/T: OR = 0.68, 95%CI, 0.48-0.95, p = 0.026). There was a significant difference in the T/A + A/A (T/A + A/A vs. T/T: OR = 0.68, 95%CI, 0.49-0.94, p = 0.020). A significant difference in T allele distribution was found in the depressed complement of SLE (T vs. A: OR = 0.67, 95%CI, 0.47-0.95, p = 0.026). There were significant differences in genetic variants of rs13137 between the positive and the negative SSB antibodies (Anti-SSB) (T vs. A: OR = 0.67, 95%CI, 0.47-0.95, p = 0.026; T/A + T/T vs. AA: OR = 2.23, 1.18-4.49, p = 0.013). The expression levels of miR-21 and miR-155 were significantly higher in patients than in controls (p < 0.001). CONCLUSIONS: This study provides novel insight that genetic variants of rs767649 and rs13137 are associated with susceptibility to SLE.

Simultaneous detection of G6PD mutations using SNPscan in a multiethnic minority area of Southwestern China.

Objectives: Baise, a multiethnic inhabited area of southwestern China, is a historical malaria-endemic area with a high prevalence of G6PD deficiency. However, few studies of G6PD deficiency have been conducted in this region. Therefore, we performed a genetic analysis of G6PD deficiency in the Baise population from January 2020 to June 2021. Methods: A SNPscan assay was developed to simultaneously detect 33 common Chinese G6PD mutations. 30 G6PD-deficient samples were used for the method's validation. Then, a total of 709 suspected G6PD-deficient samples collated from the Baise population were evaluated for G6PD status, type of mutation and effect of mutations. Results: The SNPscan test had a sensitivity of 100% [95% confidence interval (CI): 94.87%-100%] and a specificity of 100% (95% CI: 87.66%-100%) for identifying G6PD mutations. A total of fifteen mutations were identified from 76.72% (544/709) of the samples. The most common mutation was discovered to be G6PD Kaiping (24.12%), followed by G6PD Canton (17.91%), and G6PD Gaohe (11.28%). We compared the G6PD mutation spectrum among Zhuang, Han and other Southeast Asian populations, and the Zhuang population's mutation distribution was quite similar to that in the Han population. Conclusion: This study provided a detailed G6PD mutation spectrum in Baise of southwestern China and will be valuable for the diagnosis and research of G6PD deficiency in this area. Furthermore, the SNPscan assay could be used to quickly diagnose these G6PD mutations accurately.

Potential lncRNA Biomarkers for HBV-Related Hepatocellular Carcinoma Diagnosis Revealed by Analysis on Coexpression Network.

BACKGROUND: Increasing evidence demonstrated that long noncoding RNA (lncRNA) could affect inflammatory tumor immune microenvironment by modulating gene expression and could be used as a biomarker for HBC-related hepatocellular carcinoma (HCC) but still needs further research. The aim of the present study was to determine an lncRNA signature for the diagnosis of HBV-related HCC. METHODS: HBV-related HCC expression profiles (GSE55092, GSE19665, and GSE84402) were abstracted from the GEO (Gene Expression Omnibus) data resource, and R package limma and RobustRankAggreg were employed to identify common differentially expressed genes (DEGs). Using machine learning, optimal diagnostic lncRNA molecular markers for HBV-related HCC were identified. The expression of candidate lncRNAs was cross-validated in GSE121248, and an ROC (receiver operating characteristic) curve of lncRNA biomarkers was carried out. Additionally, a coexpression network and functional annotation was built, after which a PPI (protein-protein interaction) network along with module analysis were conducted with the Cytoscape open source software. RESULT: A total of 38 DElncRNAs and 543 DEmRNAs were identified with a fold change larger than 2.0 and a P value < 0.05. By machine learning, AL356056.2, AL445524.1, TRIM52-AS1, AC093642.1, EHMT2-AS1, AC003991.1, AC008040.1, LINC00844, and LINC01018 were screened out as optional diagnostic lncRNA biosignatures for HBV-related HCC. The AUC (areas under the curve) of the SVM (support vector machine) model and random forest model were 0.957 and 0.904, respectively, and the specificity and sensitivity were 95.7 and 100% and 94.3 and 86.5%, respectively. The results of functional enrichment analysis showed that the integrated coexpressed DEmRNAs shared common cascades in the p53 signaling pathway, retinol metabolism, PI3K-Akt signaling cascade, and chemical carcinogenesis. The integrated DEmRNA PPI network complex was found to be comprised of 87 nodes, and two vital modules with a high degree were selected with the MCODE app. CONCLUSION: The present study identified nine potential diagnostic biomarkers for HBV-related HCC, all of which could potentially modulated gene expression related to inflammatory conditions in the tumor immune microenvironment. The functional annotation of the target DEmRNAs yielded novel evidence in evaluating the precise functions of lncRNA in HBV-related HCC.

[Study of the genotypic and hematological feature of hemoglobin H disease in West Guangxi area].

OBJECTIVE: To analyze the incidence, genotype and hematological feature of hemoglobin H (HbH) disease in West Guangxi region. METHODS: A total of 1246 patients diagnosed with HbH disease from January 2013 to December 2018 in our hospital were enrolled. Red blood cell parameters, hemoglobin electrophoresis, Gap-polymerase china reaction (Gap-PCR) and polymerase chain reaction-reverse dot blot (PCR-RDB) techniques were used to detect the 6 common α-thalassemia mutations and 17 common ß-thalassemia mutations. The results were compared with those of other regions. RESULTS: The detection rate for HbH disease was 5.66%. Among the 1246 patients, 614 (49.28%) had deletion-type HbH disease, including -α 3.7/--SEA (35.32%),-α 4.2/--SEA(13.72%) and -a 3.7/--THAI(0.24%), 632(50.72%) had non-deleted HbH disease, mainly α CS α /--SEA (44.86%), followed by α WS α/--SEA (4.33%), α QS α /--SEA (1.45%) and α CS α/--THAI(0.08%). Co-committent HbH disease and ß-thalassemia were detected in 54 cases (4.33%). Most patients with HbH disease showed mild to moderate anemia. Very few had severe anemia. Among these, patients with HbH-CS had the most severe anemia, and HbH-WS were the mildest. Hb levels in patients with HbH disease alone were lower than those with co-committent HbH and ß-thalassemia. Compared with other regions, the incidence and genotype of HbH disease of West Guangxi are different. CONCLUSION: The prevalence of HbH disease is high in West Guangxi region, and the main genotypeis non-deletion. α CS α /--SEA is the most common, and most of them had moderate anemia. Compared with the deletion-type HbH disease, non-deleted HbH patients were more severe. When HbH disease co-committed with ß-thalassemia, the severity of anemia is reduced. The difference between West Guangxi and other regions may account for the variance of clinical manifestations and incidence of HbH disease in this region.

Serum Homocysteine Levels and Microalbuminuria in Patient with Systemic Lupus Erythematosus.

BACKGROUND: At present, the relationship between serum homocysteine and microalbuminuria (MAU) in systemic lupus erythematosus (SLE) patients is still unclear. Therefore, the aim of our study was to analyze the association between serum homocysteine and MAU in SLE patients. METHODS: The study analyzed 150 patients with SLE at Affiliated Hospital of Youjiang Medical University for Nationalities retrospectively, and we collected for clinical and laboratory data. RESULTS: We found a positive correlation between serum homocysteine and MAU in SLE patients (r = 0.430, p < 0.001). We found that serum homocysteine levels were increased in SLE patients with MAU positive compared to those who were MAU negative (p < 0.001). After adjusting for multiple confounding factors, we found that serum homocysteine maintained a positive correlation with MAU in patients with SLE in multivariate correlation analysis (p = 0.253, r = 0.002). The receiver operating characteristic (ROC) curve with an area under the curve of 0.730, and serum homocysteine had 72.2% sensitivity and 61.9% specificity with cutoff values 9.0 to identify the SLE patients with MAU positive. CONCLUSIONS: The current results found a correlation between serum homocysteine and MAU in SLE patients, suggesting that elevated serum homocysteine levels might be an adverse factor for SLE patients with kidney injury.

Exosome-transmitted long non-coding RNA SENP3-EIF4A1 suppresses the progression of hepatocellular carcinoma.

Extracellular communication mediated by exosomes in a tumor microenvironment can substantially affect tumor progression. However, the effect of exosomal long non-coding RNA SENP3-EIF4A1 on hepatocellular carcinoma (HCC) is still unclear. In this study, SENP3-EIF4A1 expressions in patients with HCC and healthy controls were detected and compared. Results showed that SENP3-EIF4A1 was significantly reduced in HCC tissues and exosomes from the plasma of patients with HCC (P<0.05) and was primarily encapsulated by exosomes. The patients with HCC and the healthy controls could be distinguished using exosomal SENP3-EIF4A1 (AUC=0.8028). The transfer of exosomal SENP3-EIF4A1 secreted by normal cells to HCC cells stimulated apoptosis and weakened the invasion and migration abilities of HCC cells to suppress their malignant biological behavior (P<0.05). Additionally, exosomal SENP3-EIF4A1 was capable of inhibiting tumor growth in vivo and modulating the expression of ZFP36 by competitively binding to miR-9-5p. In conclusion, exosomal SENP3-EIF4A1 is a new favorable biomarker for clinically detecting HCC, and SENP3-EIF4A1 can be transmitted by exosomes from normal cells to HCC cells to inhibit the in vitro and in vivo development of HCC. Thus, exosomal SENP3-EIF4A1 is involved in the communication between normal cells and HCC cells during the onset of HCC.

Long non-coding RNA F11-AS1 inhibits HBV-related hepatocellular carcinoma progression by regulating NR1I3 via binding to microRNA-211-5p.

Long non-coding RNAs (lncRNAs) could regulate growth and metastasis of hepatocellular carcinoma (HCC). In this study, we aimed to investigate the mechanism of lncRNA F11-AS1 in hepatitis B virus (HBV)-related HCC. The relation of lncRNA F11-AS1 expression in HBV-related HCC tissues to prognosis was analysed in silico. Stably HBV-expressing HepG2.2.15 cells were established to explore the regulation of lncRNA F11-AS1 by HBx protein, as well as to study the effects of overexpressed lncRNA F11-AS1 on proliferation, migration, invasion and apoptosis in vitro. Subsequently, the underlying interactions and roles of lncRNA F11-AS1/miR-211-5p/NR1I3 axis in HBV-related HCC were investigated. Additionally, the influence of lncRNA F11-AS1 and miR-211-5p on tumour growth and metastasis capacity of HepG2.2.15 cells were studied on tumour-bearing nude mice. Poor expression of lncRNA F11-AS1 was correlated with poor prognosis in patients with HBV-related HCC, and its down-regulation was caused by the HBx protein. lncRNA F11-AS1 was proved to up-regulate the NR1I3 expression by binding to miR-211-5p. Overexpression of lncRNA F11-AS1 reduced the proliferation, migration and invasion, yet induced apoptosis of HepG2.2.15 cells in vitro, which could be abolished by overexpression of miR-211-5p. Additionally, either lncRNA F11-AS1 overexpression or miR-211-5p inhibition attenuated the tumour growth and metastasis capacity of HepG2.2.15 cells in vivo. Collectively, lncRNA F11-AS1 acted as a modulator of miR-211-5p to positively regulate the expression of NR1I3, and the lncRNA F11-AS1/miR-211-5p/NR1I3 axis participated in HBV-related HCC progression via interference with the cellular physiology of HCC.

DANCR contributed to hepatocellular carcinoma malignancy via sponging miR-216a-5p and modulating KLF12.

Long noncoding RNA (lncRNA) differentiation antagonizing nonprotein coding RNA (DANCR) has been identified as an oncogene in several cancers. However, the biological function and role of DANCR in hepatocellular carcinoma (HCC) remain unclear. Our current study aimed to investigate the detailed mechanism of DANCR in HCC. We found that DANCR was significantly upregulated in HCC cell lines in comparison to LO2 cells. Then, we observed that knockdown of DANCR could greatly inhibit Huh7 and HepG2 cell proliferation. In addition, HCC cell apoptosis was increased by silence of DANCR and meanwhile, cell cycle progression was blocked in G1 phase. Apart from these, downregulation of DANCR repressed HCC cell migration and invasion ability obviously. As predicted by the bioinformatics analysis, microRNA-216a-5p (miR-216a-5p) could serve as a direct target of DANCR. MiR-216a-5p has been reported to be involved in many cancers. Here, the correlation between miR-216a-5p and DANCR was confirmed using dual-luciferase reporter assay and radioimmunoprecipitation assay. Subsequently, Kruppel-like factor 12 (KLF12) exerts an important role in different tumor types. KLF12 can function as a downstream target of miR-216a-5p. Finally, the in vivo experiments were used and the data proved that DANCR also strongly suppressed HCC tumor growth in vivo via targeting miR-216a-5p and KLF12. In conclusion, our study indicated that DANCR might provide a new perspective for HCC treatment.

Inhibition of miR-148a-3p resists hepatocellular carcinoma progress of hepatitis C virus infection through suppressing c-Jun and MAPK pathway.

OBJECTIVES: The present study was committed to investigate the role of miR-148a-3p in HCC infected with hepatitis C virus (HCV) and the regulatory mechanism of miR-148a-3p/c-Jun/MAPK signalling pathway. METHODS: Differential analysis and GSEA analysis were performed with R packages. QRT-PCR and Western blot were used to detect RNA or protein level, respectively. The targeted relationship between miR-148a-3p and c-Jun was predicted by TargetScan database and determined by double luciferase reporter assay. MTT assay and flow cytometry were used to evaluate cell proliferation, cell cycle and cell apoptosis, respectively. RESULTS: C-Jun was up-regulated, and MAPK signalling pathway was activated in HCV-infected HCC cells. C-Jun expression regulated inflammation-related gene expression and had an influence on cell proliferation, cell cycle and cell apoptosis. MiR-148a-3p, down-regulated in HCV-infected HCC cells, could target c-Jun mRNA to suppress c-Jun protein expression. CONCLUSIONS: MiR-148a-3p suppressed the proliferation of HCC cells infected with HCV through targeting c-Jun mRNA.

Antiviral effects of hepatitis B virus S gene-specific anti-gene locked nucleic acid in transgenic mice.

AIM: To assess the antiviral effects of hepatitis B virus (HBV) S gene-specific anti-gene locked nucleic acid (LNA) in transgenic mice. METHODS: Thirty HBV transgenic mice were acclimatized to laboratory conditions and positive for serum HBV surface antigen (HBsAg) and HBV DNA, were randomly divided into 5 groups (n = 7), including negative control (blank control, unrelated sequence control), positive control (lamivudine, anti-sense-LNA), and anti-gene-LNA experimental group. LNA was injected into transgenic mice by tail vein while lamivudine was administered by gavage. Serum HBV DNA and HBsAg levels were determined by fluorescence-based PCR and enzyme-linked immune sorbent assay, respectively. HBV S gene expression amounts were assessed by reverse transcription polymerase chain reaction. Positive rates of HBsAg in liver cells were evaluated immunohistochemistry. RESULTS: Average rate reductions of HBsAg after treatment on the 3rd, 5th, and 7th days were 32.34%, 45.96%, and 59.15%, respectively. The inhibitory effect of anti-gene-LNA on serum HBsAg peaked on day 7, with statistically significant differences compared with pre-treatment (0.96 ± 0.18 vs 2.35 ± 0.33, P < 0.05) and control values (P < 0.05 for all). Average reduction rates of HBV DNA on the 3rd, 5th, and 7th days were 38.55%, 50.95%, and 62.26%, respectively. This inhibitory effect peaked on the 7th day after treatment with anti-gene-LNA, with statistically significant differences compared with pre-treatment (4.17 ± 1.29 vs 11.05 ± 1.25, P < 0.05) and control values (P < 0.05 for all). The mRNA levels of the HBV S gene (P < 0.05 for all) and rates of HBsAg positive liver cells (P < 0.05 for all) were significantly reduced compared with the control groups. Liver and kidney function, and histology showed no abnormalities. CONCLUSION: Anti-gene-LNA targeting the S gene of HBV displays strong inhibitory effects on HBV in transgenic mice, providing theoretical and experimental bases for gene therapy in HBV.

Serum Bilirubin and Their Association With C-Reactive Protein in Patients With Migraine.

BACKGROUND: Increased levels of C-reactive protein (CRP) have been considered as a marker in assessing neurogenic inflammation of migraine patients. An inverse relationship between serum bilirubin and CRP has been observed in various diseases. Therefore, we analyzed serum bilirubin levels in migraine patients, and investigated the relationship between serum bilirubin and CRP in migraineurs. METHODS: A total of 86 newly diagnosed migraine patients were consecutively recruited to this study. RESULTS: Significantly lower median serum total bilirubin, conjugated bilirubin (CB) and unconjugated bilirubin were found in patients with migraine than healthy controls, and the levels of CRP were significantly higher in migraine patients than healthy controls. A negative correlation between CRP and CB was observed in patients with migraine (r = -0.255, P = 0.018). In a multiple linear regression model, the concentrations of CRP remained negatively correlated with CB. CONCLUSIONS: Our study demonstrates that serum bilirubin concentrations are decreased in migraineurs, and CB levels were found to be positively correlated with CRP in migraine patents. However, larger cross-sectional and prospective studies are needed to establish whether serum bilirubin may be a useful biomarker for assessing neurogenic inflammation in migraine patients and eventually guiding the therapy.