MicroRNA-598 inhibition ameliorates LPS-induced acute lung injury in mice through upregulating Ebf1 expression.

Acute lung injury is a critical acute respiratory distress syndrome (ARDS) with high morbidity and mortality. MicroRNAs (miRNAs) have been demonstrated to play important roles regulating acute lung injury development. In this study, we found that the expression of miR-598 was significantly upregulated in the lung tissues of mice with lipopolysaccharide (LPS)-induced acute lung injury. Both loss-of-function and gain-of-function studies were performed to evaluate the function of miR-598 in acute lung injury. The results showed that inhibition of miR-598 attenuated inflammatory response, oxidative stress, and lung injury in mice treated with LPS, while overexpression of miR-598 exacerbated the LPS-induced acute lung injury. Mechanistically, transcription factor Early B-cell Factor-1 (Ebf1) was predicted and validated as a downstream target of miR-598. Overexpression of Ebf1 attenuated LPS-induced production of inflammatory cytokine TNF-α and IL-6, ameliorated LPS-induced oxidative stress, promoted proliferation, and inhibited apoptosis in murine lung epithelial-15 (MLE-15) cells. Moreover, we demonstrated that Ebf1 knockdown abolished the protective effect of miR-598 inhibition in LPS-treated MLE-15 cells. In summary, miR-598 inhibition ameliorates LPS-induced acute lung injury in mice through upregulating Ebf1 expression, which might provide potential therapeutic treatment for acute lung injury.

Identification of sacrococcygeal and pelvic abscesses infected with invasive Mycoplasma hominis by MALDI-TOF MS.

BACKGROUND: Mycoplasma hominis is the smallest prokaryotic microorganism with no cell wall, high pleomorphism, and slower reproduction than bacteria. It is difficult for clinical technicians to find M. hominis through the negative Gram staining of specimens. Therefore, it is likely to miss detection in routine clinical smear etiological examination. M. hominis is generally considered to be a common colonizing bacterium in urogenital tract with low pathogenicity, and it is usually difficult to invade submucosal tissue and enter the bloodstream. METHODS: The abscesses of the patient were examined histopathologically, and the pus in the abscesses was extracted for etiological examination. MALDI-TOF MS was used to identify and confirmed the pathogens in the specimens. The commercial Mycoplasma isolation, culture, and drug sensitivity kit was used to determine antibiotic susceptibility. RESULTS: No pathogens were found after pathological and smear microscopic examination of the puncture fluid from the sacrococcygeal and pelvic abscesses. Until 48 h later, small, translucent, and gray-white colonies were observed in the blood plate culture results. The laboratory physician ultimately determined that the pathogen was M. hominis by MALDI-TOF MS. CONCLUSION: We report a case of extra-urogenital cystic abscesses infected by M. hominis, in order to improve clinicians' comprehensive understanding of the pathogenicity of Mycoplasma. In addition, the clinical laboratory technician should pay attention to the role of Wright-Giemsa staining of puncture fluid smear in the preliminary detection and the application of MALDI-TOF MS in identification of uncommon pathogenic microorganisms.

Variable clinical characteristics and laboratory results in five patients with Chinese Good's syndrome (thymoma and hypogammaglobulinemia): an 8-year retrospective analysis in a university hospital in China.

BACKGROUND: Good's syndrome (GS) is a rare secondary immunodeficiency disease presenting as thymoma and hypogammaglobulinemia. Due to its rarity, the diagnosis of GS is often missed. METHODS: We used the hospital information system to retrospectively screen thymoma and hypogammaglobulinemia patients at the First Affiliated Hospital of Wenzhou Medical University from Apr 2012 to Apr 2020. The clinical, laboratory, treatment, and outcome data for these patients were collected and analyzed. RESULTS: Among the 181 screened thymoma patients, 5 thymoma patients with hypogammaglobulinemia were identified; 3 patients had confirmed diagnoses of GS, and the other 2 did not have a diagnosis of GS recorded in the hospital information system. A retrospective review of the clinical characteristics, laboratory results, and follow-up data for these 2 undiagnosed patients confirmed the diagnosis of GS. All 5 GS patients presented with pneumonia, 2 patients presented with recurrent skin abscesses, 2 patients presented with recurrent cough and expectoration, 1 patient presented with recurrent oral lichen planus and diarrhea, and 1 patient presented with tuberculosis and granulomatous epididymitis. In the years after the diagnosis of hypogammaglobulinemia with mild symptoms, all 5 patients had received irregular intravenous immunoglobulin (IVIG) treatment. As the course of the disease progressed, the clinical symptoms of all patients worsened, but the symptoms were partly resolved with IVIG in these patients. However, 4 patients died due to comorbidities. CONCLUSION: GS should be investigated as a possible diagnosis in thymoma patients who present with hypogammaglobulinemia, especially those with recurrent opportunistic infections, recurrent skin abscesses, chronic diarrhea, or recurrent lichen planus.

CXCL8, CXCL9, and CXCL10 serum levels increase in syphilitic patients with seroresistance.

BACKGROUND: Recently, the rise of syphilitic seroresistance brings great confusion to the clinical diagnosis and treatment of syphilis, and no clear diagnostic marker has been found to distinguish syphilitic seroresistance from other progression of syphilis. This study evaluated the serum chemokines levels of CCL2, CXCL8, CXCL9, and CXCL10 and its correlation with blood routine, coagulation, and biochemical indexes in seroresistant syphilitic patients. METHOD: Serum levels of chemokines were quantitatively determined by Flow Cytometric Bead Array (CBA). The results expressed in pg/ml. Clinical parameters were detected and analyzed according to the clinical laboratory standards. A correlation analysis was subsequently performed. RESULTS: The seroresistant syphilitic patients increased significantly serum chemokines levels of CXCL8 (***p < 0.001), CXCL9 (***p < 0.001), and CXCL10 (**p < 0.01) when compared to noninfected individuals, but the CCL2 was not statistically significant, and serum CXCL8 shows a strong association with platelets (r = 0.51, **p = 0.004) and serum CXCL10 was significantly positively related to INR levels (r = 0.49, **p = 0.007). CONCLUSION: Increasing serum abnormalities in CXCL8, CXCL9, and CXCL10 level combining with platelets of peripheral blood and plasmatic INR in syphilis patients may be helpful for the diagnosis of serofast state.

The epidemiology and clinical features of selective immunoglobulin M deficiency: A single-center study in China.

BACKGROUND: Selective immunoglobulin M deficiency (SIgMD) is a rare primary immunodeficiency that is frequently reported in Western countries. However, large epidemiological and clinical studies of SIgMD in China are still lacking. Herein, we describe a cohort of SIgMD subjects in a large tertiary university hospital in China. METHODS: A cross-sectional study included 139 668 participants at First Affiliated Hospital of Wenzhou Medical University from January 2014 to October 2018 was conducted. Individuals with a serum IgM level less than 0.3 g/L with normal levels of serum IgA and IgG were defined as having SIgMD. RESULT: A total of 63 subjects met the criteria for SIgMD(63/139668, 0.045%), with a male-to-female ratio of 0.85, aged from 19 to 99 years. The most common clinical manifestation was autoimmune disorders (38/63, 60.32%), while the second most common manifestation was infections (21/63, 33.33%). Neither allergies nor tumors were found among these 63 SIgMD subjects. Most importantly, there were 30 patients with systemic lupus erythematosus among these 63 SIgMD subjects, accounting for 47.62% of all SIgMD subjects. CONCLUSION: To our knowledge, we describe here the first large single-center cohort of adult patients affected by SIgMD in China. The most common clinical manifestation was autoimmune disorders, specifically systemic lupus erythematosus.

The epidemiology and clinical feature of selective immunoglobulin a deficiency of Zhejiang Province in China.

BACKGROUND: Selective immunoglobulin A deficiency (SIgAD) is the most common primary antibody deficiency disease and frequently reported in the Western countries. However, large-scale epidemiologic studies on SIgAD in China are still lacking. METHODS: The clinical information of 555 180 subjects (age >4 years) including the outpatient, inpatient, and healthy subjects who had ordered serum immunoglobulin A, G, M in 9 hospitals of Zhejiang Province in China was collected. The SIgAD individuals were defined as IgA level <0.07 g/L with normal levels of serum IgG and IgM, whose age should be over 4 years, and any other secondary diseases causing SIgAD were also excluded. Then, the geographical and prevalence distribution of SIgAD individuals in Zhejiang Province and patients' clinical characteristics at the time of diagnosis were also reviewed. RESULT: Among these 555 180 subjects who had ordered the immunoglobulin evaluation, the prevalence of SIgAD was 109/555180 (0.02%). The ratio of male to female of these SIgAD individuals was 1:1.37, which also included 87 adults (≥18 years) and 22 children (18 > age >4 years). For adults, the common clinical features were infections (43/87, 49.43%), autoimmune disorders (31/87, 35.63%), allergic cases (5/87, 5.75%), and tumor cases (4/87, 4.60%). Additionally, infectious diseases (20/22, 90.91%), autoimmune disorders (4/22, 18.18%), and allergic cases (1/22, 4.55%) were found in 22 children. CONCLUSION: We first describe a large cohort of SIgAD individuals of Zhejiang Province in China. The incidence was 0.020%. The common clinical features were infection, autoimmune disorders, tumor, and allergy, and the infection rate was higher in children than the adults.

Functional screen reveals essential roles of miR-27a/24 in differentiation of embryonic stem cells.

MicroRNAs play important roles in controlling the embryonic stem cell (ESC) state. Although much is known about microRNAs maintaining ESC state, microRNAs that are responsible for promoting ESC differentiation are less reported. Here, by screening 40 microRNAs pre-selected by their expression patterns and predicted targets in Dgcr8-null ESCs, we identify 14 novel differentiation-associated microRNAs. Among them, miR-27a and miR-24, restrained by c-Myc in ESC, exert their roles of silencing self-renewal through directly targeting several important pluripotency-associated factors, such as Oct4, Foxo1 and Smads. CRISPR/Cas9-mediated knockout of all miR-27/24 in ESCs leads to serious deficiency in ESC differentiation in vitro and in vivo. Moreover, depleting of them in mouse embryonic fibroblasts can evidently promote somatic cell reprogramming. Altogether, our findings uncover the essential role of miR-27 and miR-24 in ESC differentiation and also demonstrate novel microRNAs responsible for ESC differentiation.

Diagnosing malignant pleural effusion using clinical and analytical parameters.

BACKGROUND: Malignant pleural effusion (MPE) is common and diagnosis is often problematic. A cancer ratio (serum lactate dehydrogenases: pleural adenosine deaminase ratio) has been proposed for diagnosing MPE. However, the usefulness of this "cancer ratio" and the clinical-radiological criteria for diagnosing MPE has not been clearly determined to date. The aim of this study was to assess the performance of those parameters in the diagnosis of MPE. METHODS: We analyzed 240 patients including 120 with MPE and 120 with non-MPE (93 tuberculous and 27 parapneumonic). Patients were divided into two groups: MPE and non-MPE (eg, tuberculous and parapneumonic). We constructed two predictive models to assess the probability of MPE: (a) clinical-radiological data only and (b) a combination of clinical-radiological data, the cancer ratio, and the carcinoembryonic antigen (CEA). The performances of the predictive models were assessed using receiver operating characteristic (ROC) curves and by examining the calibration. RESULTS: The area under the ROC curves for model 1 and model 2 were excellent, 0.936 and 0.998, respectively. The overall diagnostic accuracies for model 1 and model 2 were 87.5% and 98.8%, respectively. CONCLUSION: The results confirm that both models achieved a high diagnostic accuracy for MPE; however, model 2 was superior with the addition of its simplicity of use in daily practice. This model should be applied to determine which patients with a pleural effusion of unknown origin would not benefit from further invasive procedures.

The PU.1-Modulated MicroRNA-22 Is a Regulator of Monocyte/Macrophage Differentiation and Acute Myeloid Leukemia.

MicroRNA-22 (miR-22) is emerging as a critical regulator in organ development and various cancers. However, its role in normal hematopoiesis and leukaemogenesis remains unclear. Here, we detected its increased expression during monocyte/macrophage differentiation of HL-60, THP1 cells and CD34+ hematopoietic stem/progenitor cells, and confirmed that PU.1, a key transcriptional factor for monocyte/macrophage differentiation, is responsible for transcriptional activation of miR-22 during the differentiation. By gain- and loss-of-function experiments, we demonstrated that miR-22 promoted monocyte/macrophage differentiation, and MECOM (EVI1) mRNA is a direct target of miR-22 and MECOM (EVI1) functions as a negative regulator in the differentiation. The miR-22-mediated MECOM degradation increased c-Jun but decreased GATA2 expression, which results in increased interaction between c-Jun and PU.1 via increasing c-Jun levels and relief of MECOM- and GATA2-mediated interference in the interaction, and thus promoting monocyte/macrophage differentiation. We also observed significantly down-regulation of PU.1 and miR-22 as well as significantly up-regulation of MECOM in acute myeloid leukemia (AML) patients. Reintroduction of miR-22 relieved the differentiation blockage and inhibited the growth of bone marrow blasts of AML patients. Our results revealed new function and mechanism of miR-22 in normal hematopoiesis and AML development and demonstrated its potential value in AML diagnosis and therapy.

A regulatory circuit comprising GATA1/2 switch and microRNA-27a/24 promotes erythropoiesis.

Transcriptional networks orchestrate complex developmental processes, and such networks are commonly instigated by master regulators for development. By now, considerable progress has been made in elucidating GATA factor-dependent genetic networks that control red blood cell development. Here we reported that GATA-1 and GATA-2 co-regulated the expression of two microRNA genes, microRNA-27a and microRNA-24, with critical roles in regulating erythroid differentiation. In general, GATA-2 occupied the miR-27a≈24 promoter and repressed their transcription in immature erythroid progenitor cells. As erythropoiesis proceeded, GATA-1 directly activated miR-27a≈24 transcription, and this involved a GATA-1-mediated displacement of GATA-2 from chromatin, a process termed 'GATA switch'. Furthermore, the mature miR-27a and miR-24 cooperatively inhibited GATA-2 translation and favoured the occupancy switch from GATA-2 to GATA-1, thus completing a positive feedback loop to promote erythroid maturation. In line with the essential role of GATA factors, ectopic expression of miR-27a or miR-24 promoted erythropoiesis in human primary CD34+ haematopoietic progenitor cells and mice, whereas attenuated miR-27 or miR-24 level led to impaired erythroid phenotypes in haematopoietic progenitor cells and zebrafish. Taken together, these data integrated micro RNA expression and function into GATA factor coordinated networks and provided mechanistic insight into a regulatory circuit that comprised GATA1/2 switch and miR-27a/24 in erythropoiesis.

The regulatory roles of microRNA-146b-5p and its target platelet-derived growth factor receptor α (PDGFRA) in erythropoiesis and megakaryocytopoiesis.

Emerging evidence has shown that microRNAs have key roles in regulating various normal physiological processes, whereas their deregulated expression is correlated with various diseases. The miR-146 family includes miR-146a and miR-146b, with a distinct expression spectrum in different hematopoietic cells. Recent work indicated that miR-146a has a close relationship with inflammation and autoimmune diseases. miR-146-deficient mice have developed some abnormal hematopoietic phenotypes, suggesting the potential functions of miR-146 in hematopoietic development. In this study, we found that miR-146b was consistently up-regulated in both K562 and CD34(+) hematopoietic stem/progenitor cells (HSPCs) undergoing either erythroid or megakaryocytic differentiation. Remarkably, erythroid and megakaryocytic maturation of K562 cells was induced by excess miR-146b but inhibited by decreased miR-146b levels. More importantly, an mRNA encoding receptor tyrosine kinase, namely platelet-derived growth factor receptor α (PDGFRA), was identified and validated as a direct target of miR-146b in hematopoietic cells. Gain-of-function and loss-of-function assays showed that PDGFRA functioned as a negative regulator in erythroid and megakaryocytic differentiation. miR-146b could ultimately affect the expression of the GATA-1 gene, which is regulated by HEY1 (Hairy/enhancer-of-split related with YRPW motif protein 1), a transcriptional repressor, via inhibition of the PDGFRA/JNK/JUN/HEY1 pathway. Lentivirus-mediated gene transfer also demonstrated that the overexpression of miR-146b promoted erythropoiesis and megakaryocytopoiesis of HSPCs via its regulation on the PDGFRA gene and effects on GATA-1 expression. Moreover, we confirmed that the binding of GATA-1 to the miR-146b promoter and induction of miR-146b during hematopoietic maturation were dependent on GATA-1. Therefore, miR-146b, PDGFRA, and GATA-1 formed a regulatory circuit to promote erythroid and megakaryocytic differentiation.

DNA methylation downregulated mir-10b acts as a tumor suppressor in gastric cancer.

BACKGROUND: MicroRNAs act as tumor suppressors or oncogenes. The pathological roles of miRNAs in gastric tumorigenesis are largely unknown. Although miR-10b was identified as an miRNA deregulator expressed in gastric cancer (GC), there also exists some debate on whether miR-10b is acting as tumor suppressor or oncogene in GC. METHODS: Quantitative RT-PCR was employed to investigate the level of miR-10b in GC tissues and matched adjacent normal tissues (n = 100). In vitro cell proliferation, apoptosis assays, cell migration, and invasion assays were performed to elucidate the biological effects of miR-10b. Because silencing of miRNA by promoter CpG island methylation may be an important mechanism in tumorigenesis, GC cells were treated with 5-aza-2'-deoxycytidine and trichostatin A, and expression changes of miR-10b were subsequently examined by quantitative RT-PCR. Furthermore, the methylation status of the CpG island upstream of miR-10b was analyzed by methylation-specific PCR in GC tissues (n = 29). RESULTS: We showed here that miR-10b was significantly downregulated in GC cell lines and tissues as demonstrated by quantitative real-time PCR. Overexpression of miR-10b in MGC-803 and HGC-27 dramatically suppressed cell proliferation, migration, invasion, and induced apoptosis. Moreover, we demonstrated that T-cell lymphoma invasion and metastasis (Tiam1) was a target of miR-10b. Furthermore, 5-aza-2'-deoxycytidine and trichostain A increased miR-10b expression, and the methylation level was high in the CpG islands upstream of miR-10b gene. CONCLUSIONS: Taken together, these findings suggest that miR-10b may function as a novel tumor suppressor and is partially silenced by DNA hypermethylation in GC.

ZNF16 (HZF1) promotes erythropoiesis and megakaryocytopoiesis via regulation of the c-KIT gene.

We previously characterized the zinc finger protein gene HZF1 [also known as ZNF16 (zinc finger protein 16)] and demonstrated its important roles in erythroid and megakaryocytic differentiation of K562 cells. In the present study, we investigated its effect on erythroid and megakaryocytic differentiation of HSPCs (haemopoietic stem/progenitor cells). We observed up-regulation of ZNF16 during erythroid and megakaryocytic differentiation of the CD34+ HSPCs, and demonstrated that ZNF16 promotes erythroid and megakaryocytic differentiation by gain-of-function and loss-of-function experiments. Using a luciferase reporter and ChIP assays ZNF16 was demonstrated to bind to the c-KIT gene promoter and inhibit its expression in K562 cells. Enforced expression and knockdown of ZNF16 down-regulated and up-regulated the expression of the c-KIT gene in K562 cells and HSPCs respectively. Significantly decreased levels of the c-Kit protein were observed following erythroid and megakaryocytic differentiation of K562 and CD34+ cells. The knockdown of c-KIT partially rescued the differentiation inhibition caused by ZNF16 knockdown. The knockdown of c-KIT also blocked the activity of the c-Raf/MEK [MAPK (mitogen-activated protein kinase)/ERK (extracellular-signal-regulated kinase) kinase]/ERK/c-Jun signal pathway and reduced further the level of HEY1 (hes-related family bHLH transcription factor with YRPW motif 1), a repressor of GATA1 (GATA-binding protein 1) transcription, which finally up-regulated the expression of GATA1, a central regulator of erythroid and megakaryocytic differentiation. In conclusion the results of the present study demonstrate that ZNF16 plays an important role in erythropoiesis and megakaryocytopoiesis via its regulation of the c-Kit/c-Raf/MEK/ERK/c-Jun/HEY1/GATA1 cascade.

A comprehensive analysis of GATA-1-regulated miRNAs reveals miR-23a to be a positive modulator of erythropoiesis.

miRNAs play important roles in many biological processes, including erythropoiesis. Although several miRNAs regulate erythroid differentiation, how the key erythroid regulator, GATA-1, directly orchestrates differentiation through miRNA pathways remains unclear. In this study, we identified miR-23a as a key regulator of erythropoiesis, which was upregulated both during erythroid differentiation and in GATA-1 gain-of-function experiments, as determined by miRNA expression profile analysis. In primary human CD34+ hematopoietic progenitor cells, miR-23a increased in a GATA-1-dependent manner during erythroid differentiation. Gain- or loss-of-function analysis of miR-23a in mice or zebrafish demonstrated that it was essential for normal morphology in terminally differentiated erythroid cells. Furthermore, a protein tyrosine phosphatase, SHP2, was identified as a downstream target of miR-23a that mediated its regulation of erythropoiesis. Taken together, our data identify a key GATA-1-miRNA axis in erythroid differentiation.

Characterization of microRNA-29 family expression and investigation of their mechanistic roles in gastric cancer.

Increasing evidence shows that abnormal microRNAs (miRNAs) expression is involved in tumorigenesis. They might be the novel biomarkers or therapeutic targets in disease treatment. miR-29 family was previously reported to act as tumor suppressors or oncogenes in diverse cancers. However, their accurate expression, function and mechanism in gastric cancer (GC) are not well known. Here, we found that the expression of miR-29 family members was significantly reduced in GC compared with adjacent controls. Among them, miR-29c had the most reduced percentage in GC and was associated with aggressive and progressive phenotypes of GC. We further demonstrated that miR-29 family acted as tumor suppressors through targeting CCND2 and matrix metalloproteinase-2 genes in GC. Moreover, the inverse relationship between miR-29 family and their targets was verified in patients and xenograft mice. Finally, reintroduction of miR-29 family significantly inhibited tumor formation of GC cells in the xenograft mice. Take together, our finding characterized the expression properties of miR-29 family, contributed to the function and molecular mechanism of miR-29 family in GC and implied that miR-29 family might be employed as novel prognostic markers and therapeutic targets of GC.

MicroRNA-29a and microRNA-142-3p are regulators of myeloid differentiation and acute myeloid leukemia.

Although microRNAs (miRNAs) are increasingly linked to various physiologic processes, including hematopoiesis, their function in the myeloid development is poorly understood. We detected up-regulation of miR-29a and miR-142-3p during myeloid differentiation in leukemia cell lines and CD34(+) hematopoietic stem/progenitor cells. By gain-of-function and loss-of-function experiments, we demonstrated that both miRNAs promote the phorbol 12-myristate 13-acetate-induced monocytic and all-trans-retinoic acid-induced granulocytic differentiation of HL-60, THP-1, or NB4 cells. Both the miRNAs directly inhibited cyclin T2 gene, preventing the release of hypophosphorylated retinoblastoma and resulting in induction of monocytic differentiation. In addition, a target of miR-29a, cyclin-dependent kinase 6 gene, and a target of miR-142-3p, TGF-ß-activated kinase 1/MAP3K7 binding protein 2 gene, are involved in the regulation of both monocytic and granulocytic differentiation. A significant decrease of miR-29a and 142-3p levels and an obvious increase in their target protein levels were also observed in blasts from acute myeloid leukemia. By lentivirus-mediated gene transfer, we demonstrated that enforced expression of either miR-29a or miR-142-3p in hematopoietic stem/progenitor cells from healthy controls and acute myeloid leukemia patients down-regulated expression of their targets and promoted myeloid differentiation. These findings confirm that miR-29a and miR-142-3p are key regulators of normal myeloid differentiation and their reduced expression is involved in acute myeloid leukemia development.