A 16-color full spectrum flow cytometric analysis for comprehensive evaluation of T-cell reconstitution in SIV-infected rhesus macaques.

T-cell reconstitution is central in human immunodeficiency virus (HIV) infection/disease progression. Simian immunodeficiency virus (SIV)-infected rhesus macaques (Macaca mulatta) have been the most widely used animal model for HIV research so far. An effective flow cytometry panel is crucial for monitoring the T cell reconstitution in SIV infection progression. We developed this sixteen-color flow cytometry-based panel for a T cell subsets analysis by manual gating and, once successfully gated, to characterize T cells function in-depth in rhesus macaques. This panel included markers to characterize CD4+ T cells and CD8+ T cells, T regulatory cells (Tregs), and T cell differentiation status (CD45RA and CCR7). Additionally, we included antibodies that measure T cell activation and proliferation molecules (CD69, HLA-DR, CD38 and Ki67), antibodies that examine the expressions of key PD-1 pathway molecule (PD-1), SIV potential target (CD32) and the primary SIV co-receptor CCR5 (CD195). High-dimensional single cell analysis was also performed to identify CD3+ T cells immunophenotypes of SIV-infected rhesus macaques. We designed this panel to evaluate the responses of different T cell subsets to SIV in whole blood from SIV-infected rhesus macaques.

Exposure to Ambient Air Pollutant PM10 in the Second Trimester of Pregnancy Is Associated with Preterm Birth: A Birth-Based Health Information Cohort Study.

Objectives: We evaluated the effects of exposure to high concentrations of particulate matter (PM)10 on preterm birth (PTB) and identified a critical concentration of PM10 that could lead to PTB via a birth-based health information cohort study. Methods: We conducted a birth-based cohort study consisting of nonanomalous singleton births at 22-42 weeks. PTB was defined as babies born alive before 37 weeks of pregnancy. Pregnancy period exposure averages were estimated for PM10 based on the China National Environmental Monitoring Centre (CNEMC). Pregnant women who lived within 50 km of the monitor station were recruited into this study. Logistic regression analyses were performed to determine the association between PTB and exposure to PM10 at different pregnancy periods with adjustment for confounding factors. Results: The relative frequency of PTB was 8.7% in the study cohort of 5,291 singleton live births. A total of 1137 women had a high level of PM10 exposure (≥60 µg/m3) in the second trimester of pregnancy. The average concentrations of PM10 in the first, second, and third trimesters of pregnancy and throughout pregnancy were 53.8 µg/m3, 54.2 µg/m3, 55.6 µg/m3, and 54.3 µg/m3, respectively. The generalized additive model (GAM) analysis showed that there was a nonlinear correlation between PM10 and PTB in the second trimester of pregnancy (P < 0.001). The adjusted odds ratio between PTB and low concentration PM10 exposure (PM10 < 60 µg/m3) in the second trimester of pregnancy was 1.01 (95% CI 0.95-1.05). However, high PM10 exposure (PM10 ≥ 60 µg/m3) in the second trimester of pregnancy had an increased PTB risk even after adjustment for coexisting risk factors with an adjusted odds ratio of 1.78 (95% CI 1.69-1.87), and the incidence of PTB increased with an increase in PM10 exposure. Conclusions: Our research discovered that exposure to high levels of PM10 increases the risk of PTB and the second trimester is the most vulnerable gestational period to ambient air pollution exposure. PM10 concentrations more than 60 µg/m3 are detrimental to pregnant women in their second trimester. This study has implications for health informatics-oriented healthcare decision support systems.

VPS9D1-AS1, a novel long-non-coding RNA, acts as a tumor promoter by regulating the miR-324-5p/ITGA2 axis in colon adenocarcinoma.

BACKGROUND: Colon adenocarcinoma (COAD) is among the most common malignancies worldwide. Elucidating the function and mechanism of action of the lncRNA VPS9D1-AS1 in COAD will be of great value for identifying potential therapeutic targets. METHODS: Quantitative real-time polymerase chain reaction (qRT-PCR) analysis was used to measure the expression levels of lncRNA VPS9D1-AS1 in COAD tissues and cell lines. After knocking down the expression of VPS9D1-AS1 in two COAD cell lines, namely SW1116 and LoVo, their proliferation rate was measured by the 5-ethynyl-2'-deoxyuridine (Edu) incorporation and cell counting kit-8 (CCK-8) viability assays, migration and invasion abilities were assessed by wound healing and Transwell assays, and apoptosis rate was measured withflow cytometry. Additionally, the dual luciferase reporter assay system was used to investigate the targeting of miR-324-5p to VPS9D1-AS1 and ITGA2 3'-UTR. The inhibitory effects of the miR-324-5p/ITGA2 axis on the function of VPS9D1-AS1 were also examined. In vivo tumorigenesis assay was performed in nude mice injected with VPS9D1-AS1 shRNA or control shRNA lentivirus-transfected LoVo cells. RESULTS: VPS9D1-AS1 was found to be upregulated in COAD tissues and cell lines. VPS9D1-AS1 knockdown inhibited the COAD cell proliferation, migration and invasion and increased the apoptosis rate. In addition, we have demonstrated that miR-324-5p targets VPS9D1-AS1 and ITGA2 3'-UTR, and miR-324-5p silencing or forced ITGA2 expression attenuated the effect of VPS9D1-AS1 knockdown. CONCLUSION: This study identified a novel competing endogenous RNA (ceRNA) pathway that potentially associates with the oncogenic functions of VPS9D1-AS1, miR-324-5p, and ITGA2 in COAD, which could contribute to the identification of new therapeutic approaches targeting COAD.

Comparison of clinical and imaging features between pulmonary tuberculosis complicated with lung cancer and simple pulmonary tuberculosis: a systematic review and meta-analysis.

This review aimed to compare the clinical features and CT imaging features between patients with pulmonary tuberculosis (PTB) and lung cancer and patients with PTB alone. That would help to analyse the differences between the two and consequently providing a theoretical basis for the clinical diagnosis and treatment for the patients. Relevant case-control studies focusing on the clinical and CT imaging characteristics between PTB with lung cancer and PTB alone were systematically searched from five electronic databases. Odds ratios (ORs) and 95% confidence intervals (CIs) were calculated for comparison. As of 2021-07-06, a total of 1735 articles were retrieved. But only 15 articles were finally included for meta-analysis. The results showed a higher proportion of irritable cough, haemorrhagic pleural effusion and lower proportion of night sweating in PTB patients with lung cancer than in PTB patients, and the differences were statistically significant (irritable cough: OR 2.43, 95% CI 1.43-4.11; haemorrhagic pleural effusion: OR 5.73, 95% CI 1.63-20.12; night sweating: OR 0.56, 95% CI 0.36-0.87). In addition, there are many differences in the imaging characteristics of the two types of patients. In conclusion, this review summarises the similarities and differences in clinical symptoms and imaging features between patients with PTB and lung cancer and patients with PTB alone, suggesting that we should be alert to the occurrence of lung cancer in patients with obsolete PTB relapse.

miR-23b-3p Inhibits the Oncogenicity of Colon Adenocarcinoma by Directly Targeting NFE2L3.

BACKGROUND AND AIMS: MicroR-23b-3p (miR-23b-3p) has been found to be abnormally expressed in a variety of malignant tumors and to play a role in tumor inhibition or promotion. However, the regulatory mechanism of miR-23b-3p in COAD remains unclear. The purpose of this study was to investigate the clinical significance of miR-23b-3p expression in COAD cells and to explore its role and regulatory mechanism in the growth of COAD. MATERIALS AND METHODS: Quantitative real-time polymerase chain reaction (qRT-PCR) was used to measure miR-23b-3p expression in COAD tissues and cell lines. After transfecting miR-23b-3p mimics into two human COAD cell lines (SW620 and LoVo), the cell counting kit-8 (CCK-8), colony formation, and 5-ethynyl-2'-deoxyuridine (EdU) assays were used to detect cell proliferation, the Transwell assay was used to measure cell migration and invasion capacity, and flow cytometry was used to evaluate cell apoptosis in vitro. In addition, a luciferase reporter assay was used to determine whether miR-23b-3p targets NFE2L3. The downstream regulatory mechanisms of miR-23b-3p action in COAD cells were also investigated. For in vivo tumorigenesis assay, COAD cells stably overexpressing miR-23b-3p were injected subcutaneously into the flank of nude mice to obtain tumors. RESULTS: Significantly decreased expression of miR-23b-3p was detected in COAD tissues and cell lines. Exogenous miR-23b-3p expression inhibited cell proliferation, migration, and invasion and promoted cell apoptosis of COAD cells in vitro. Nuclear factor erythroid 2 like 3 (NFE2L3) was identified as a direct target gene of miR-23b-3p. In addition, reintroduction of NFE2L3 partially abolished the anticancer effects of miR-23b-3p on COAD cells. Furthermore, miR-23b-3p overexpression hindered the growth of COAD cells in vivo. CONCLUSION: miR-23b-3p inhibited the oncogenicity of COAD cells in vitro and in vivo by directly targeting NFE2L3, suggesting the importance of the miR-23b-3p/NFE2L3 pathway in the development of COAD.

Prenatal diagnosis of Cri-du-Chat syndrome with concomitant distal trisomy 10q syndrome in one fetus with ultrasound anomalies.

OBJECTIVE: The aim of this work was to characterize the genetic abnormalities and prenatal diagnosis indications in one fetus with Cri-du-Chat syndrome with codependent 10q24.2-q26.3 duplication in prenatal screening. MATERIALS AND METHODS: A 31-year-old woman had a second trimester serum screening that indicated the fetus was at low risk. During this pregnancy, the woman underwent amniocentesis at 18+4 weeks' gestation because of adverse fertility history and nuchal fold thickening. Cytogenetic analysis and next-generation sequencing analysis were simultaneously performed to provide genetic analysis of fetal amniotic fluid. According to abnormal results, parental chromosome karyotype of peripheral blood was performed to analysis. RESULTS: CNV-seq detected a 14.00 Mb deletion at 5p15.33-p15.2 and a 34.06 Mb duplication at 10q24.2-q26.3 in the fetus. Cytogenetic analysis of the fetus revealed a karyotype of 46, XY, der(5) t(5;10) (p15.2;q26.3). The karyotype of pregnant women was 46,XX,t(5;10) (p15.2;q24.2). The pregnancy was subsequently terminated after sufficient informed consent. CONCLUSION: This is the first study that reports prenatal diagnosis of a Cri-du-Chat syndrome with concomitant 10 q24.2-q26.3 duplication. Adverse pregnancy history has to be as an important indicator for prenatal diagnosis, and the genetic factors of abnormal pregnancy should be identified before next pregnancy. Nuchal fold thickening is closely related to fetal abnormalities. Combined with ultrasonography, the use of CNV-seq will improve the diagnosis of submicroscopic chromosomal aberrations in fetuses with congenital anomalies.

Novel lncRNA SFTA1P Promotes Tumor Growth by Down-Regulating miR-4766-5p via PI3K/AKT/mTOR Signaling Pathway in Hepatocellular Carcinoma.

BACKGROUND: Hepatocellular carcinoma (HCC) is a common malignancy worldwide with a high mortality rate. lncRNA SFTA1P is highly expressed in HCC. We aimed to study the role of SFTA1P in HCC and its relationship with miR-4766-5p. MATERIALS AND METHODS: The levels of SFTA1P in HCC tissues and cell lines were determined. Relationship between SFTA1P and clinical features and prognosis was studied. The influence of SFTA1P on HCC cell viability, migration, invasion and apoptosis was studied in vitro. Rescue experiments were conducted after the binding site between SFTA1P and miR-4766-5p confirmed by dual-luciferase assay. The protein expression of AKT, p-AKT, mTOR and p-mTOR in HCC cells with knockdown of SFTA1P was determined by Western blotting. A tumor study in nude mice was conducted in order to assess the effects of SFTA1P on tumor growth characteristics. RESULTS: SFTA1P was up-regulated in HCC tissues and cell lines. SFTA1P expression was closely related to tumor size, vascular invasion and TNM stage. Knockdown of SFTA1P inhibited HCC cell viability, migration and invasion and promoted cell apoptosis. MiR-4766-5p was a target of SFTA1P and knockdown of SFTA1P could decrease the protein expression of p-AKT and p-mTOR. Rescue experiments showed that miR-4766-5p mimics could attenuate the promoting role of SFTA1P on HCC cell viability, invasion and migration, and inhibiting role on cell apoptosis. Moreover, we used nude mice models and also found that the knockdown of SFTA1P reduced tumor volume and weight. CONCLUSION: lncRNA SFTA1P could promote tumor development in HCC by down-regulating miR-4766-5p expression via PI3K/AKT/mTOR signaling pathway. It may be a potential therapeutic target for HCC.

Chronic psychological stress impairs germinal center response by repressing miR-155.

Germinal centers (GC) are vital to adaptive immunity. BCL6 and miR-155 are implicated in control of GC reaction and lymphomagenesis. FBXO11 causes BCL6 degradation through ubiquitination in B-cell lymphomas. Chronic psychological stress is known to drive immunosuppression. Corticosterone (CORT) is an adrenal hormone expressed in response to stress and can similarly impair immune functions. However, whether GC formation is disrupted by chronic psychological stress and its molecular mechanism remain to be elucidated. To address this issue, we established a GC formation model in vivo, and a GC B cell differentiation model in vitro. Comparing Naive B cells to GC B cells in vivo and in vitro, the differences of BCL6 and FBXO11 mRNA do not match the changes at the protein level and miR-155 levels that were observed. Next we demonstrated that CORT increase, induced by chronic psychological stress, reduced GC response, IgG1 antibody production and miR-155 level in vivo. The effect of chronic psychological stress can be blocked by a glucocorticoid receptor (GR) antagonist. Similarly, impaired GC B cell generation and isotope class switching were observed. Furthermore, we found that miR-155 and BCL6 expression were downregulated, but FBXO11 expression was upregulated in GC B cells treated with CORT in vitro. In addition, we demonstrated that miR-155 directly down-regulated FBXO11 expression by binding to its 3́-untranslated region. The subsequent overexpression of miR-155 significantly blocked the stress-induced impairment of GC response, due to changes in FBXO11 and BCL6 expression, as well as increased apoptosis in B cells both in vivo and in vitro. Our findings suggest perturbation of GC reaction may play a role in chronic psychological stress-induced immunosuppression through a glucocorticoid pathway, and miR-155-mediated post-transcriptional regulation of FBXO11 and BCL6 expression may contribute to the impaired GC response.

[A preliminary study on immune adjunctive therapy for multidrug-resistant Mycobacterium tuberculosis infection in animal models].

OBJECTIVE: To investigate the effect of adjunctive therapy by immune agents in mice infected with multidrug-resistant tuberculosis (MDR-TB). METHODS: Sixty-eight adult male BALB/c mice were infected with multidrug-resistant Mycobacterium tuberculosis (MTB) by aerosol route. The mice were randomly divided into a control group, an immuno-treatment group, a drug treatment group and a combination treatment group (drug plus immuno-treatment). In each treatment group, 16 mice were treated at day 21 after infection, and another 4 mice were sacrificed at day 21 after infection (treatment for 0 week) as the blank control group. In the treatment group, 4 mice were sacrificed in turn at the day after treatment for 4, 8, 16 and 20 weeks. Lung and spleen mass index at the day after treatment for 8, 16 and 20 weeks, lung and spleen live bacterial count in each period, serum IFN-γ and IL-10 levels at the day after treatment for 8 weeks were measured. Comparisons of analyzed parameters among groups were performed with the one way ANOVA test, and comparisons of parameters between 2 groups were performed with SNK and Games-Howell test. RESULTS: The lung mass index in the immuno-treatment group (0.66 ± 0.09)%, drug group (0.60 ± 0.07)% and combination therapy group (0.57 ± 0.05)% at the day after treatment for 8 weeks were significantly lower than that of the control group (0.81 ± 0.09)%, (F = 7.364, P < 0.01). Spleen CFU of immuno-treatment group at 16 and 20 weeks [(3.11 ± 0.14) lg CFU and (3.15 ± 0.18) lg CFU] were significantly lower than those of the control group [(3.77 ± 0.35) lg CFU and (4.31 ± 0.06)] (F values were 446.424 and 2107.689, P < 0.01). Spleen tissues of the drug group and the combination therapy group were sterile from 4 weeks. The serum IFN-γ levels of immuno-treatment group, drug group and combination therapy were (5.3 ± 1.9) ng/L, (1.3 ± 0.5) ng/L and (0.9 ± 1.3) ng/L, respectively, being significantly lower than that of the control group (10.3 ± 2.1) ng/L (F = 32.128, P < 0.01). The lung and spleen mass index, lung and spleen CFU, serum IFN-γ and IL-10 between medication group and combination therapy showed no significant differences. CONCLUSIONS: Immuno-treatment could mitigate lung tissue inflammation, reduce the number of MTB in mouse spleen tissues and decrease serum IFN-γ levels in the MDR-TB mouse model. However immuno-treatment failed to reduce the number of MTB in lung tissues. There was no adjunctive effect of immuno treatment for MDR-TB mice in reducing the number of MTB and mitigating inflammation.