Impaired thymic export and apoptosis contribute to regulatory T-cell defects in patients with chronic heart failure

Objective

Animal studies suggest that regulatory T (T_reg) cells play a beneficial role in ventricular remodeling and our previous data have demonstrated defects of T_reg cells in patients with chronic heart failure (CHF). However, the mechanisms behind T_reg-cell defects remained unknown. We here sought to elucidate the mechanism of T_reg-cell defects in CHF patients.

Methods and Results

We performed flow cytometry analysis and demonstrated reduced numbers of peripheral blood CD4⁺CD25⁺FOXP3⁺CD45RO⁻CD45RA⁺ naïve T_reg (nT_reg) cells and CD4⁺CD25⁺FOXP3⁺CD45RO⁺CD45RA⁻ memory T_reg (mT_reg) cells in CHF patients as compared with non-CHF controls. Moreover, the nT_reg/mT_reg ratio (p<0.01), CD4⁺CD25⁺FOXP3⁺CD45RO⁻ CD45RA⁺CD31⁺ recent thymic emigrant T_reg cell (RTE-T_reg) frequency (p<0.01), and T-cell receptor excision circle levels in T_reg cells (p<0.01) were lower in CHF patients than in non-CHF controls. Combined annexin-V and 7-AAD staining showed that peripheral T_reg cells from CHF patients exhibited increased spontaneous apoptosis and were more prone to interleukin (IL)-2 deprivation- and CD95 ligand-mediated apoptosis than those from non-CHF individuals. Furthermore, analyses by both flow cytometry and real-time polymerase chain reaction showed that T_reg-cell frequency in the mediastinal lymph nodes or Foxp3 expression in hearts of CHF patients was no higher than that of the non-CHF controls.

Conclusion

Our data suggested that the T_reg-cell defects of CHF patients were likely caused by decreased thymic output of nascent T_reg cells and increased susceptibility to apoptosis in the periphery.     

玉米单染色体的分离和体外扩增

建立了玉米单染色体的分离及体外扩增的方法。取95％乙醇固定后经果胶酶和纤维酶酶解的根尖制备染色体标本,用自制的微细玻璃针在倒置显微镜下挑取目的染色体。染色体DNA经Sau3A酶切后与人工合成的Sau3A连接接头连接,经两次PCR扩增获得足以用于构建单染色体DNA文库的扩增产物。片段大小为0．3～5kb,多数为0．5～3．5kb．与前人研究方法相比,所需底物量少（只需1条染色体）,扩增片段大,为植物中小型染色体分离、体外扩增进而进行单染色体DNA文库构建奠定了基础。     

低温条件下广聚萤叶甲成虫的耐饥饿能力

【背景】广聚萤叶甲是恶性入侵杂草豚草的一种重要专一性天敌,在冬季低温且缺乏食物的条件下,广聚萤叶甲的耐饥饿能力直接关系到其越冬种群的虫源基数。【方法】在室内观察广聚萤叶甲成虫在10℃低温条件下的存活率和死亡率,研究了水分(有水、无水)、不同密度(一雌一雄、二雌二雄、五雌五雄)、加入泥土枯枝对广聚萤叶甲耐饥饿能力的影响。【结果】无水、有水条件下,广聚萤叶甲成虫的平均存活时间分别为(15.23±1.01)、(13.33±0.88)d,水分对广聚萤叶甲的耐饥力的影响不显著;随着密度的增加,广聚萤叶甲的耐饥饿能力增强,一雌一雄、二雌二雄和五雌五雄的平均存活时间依次为(11.96±0.57)、(13.78±0.60)、(14.81±0.42)d;加入泥土和豚草枯枝后,广聚萤叶甲的耐饥饿能力明显提高,其平均存活时间为(15.97±1.05)d。【结论与意义】低温条件下广聚萤叶甲的耐饥饿能力较强,可保证部分广聚萤叶甲的自然种群在野外安全越冬,研究低温条件下广聚萤叶甲的耐饥饿能力对豚草的生物防治具有重要意义。     

鼠伤寒沙门氏菌SL7207 SifA-突变株的构建和鉴定

鼠伤寒沙门氏菌SifA^-基因突变株的特点是能进入真核细胞的胞液。利用P22噬菌体转导技术构建了鼠伤寒沙门氏菌疫苗株SL7207的SifA^-突变株SL7207,该突变株与SL7207有着相似的体外生长曲线和细胞侵袭力,SL7207。在MDCK上皮细胞中的增殖能力增强,但在RAW264．7巨噬细胞中的生存能力减弱。小鼠毒力试验显示SL7207。在BALB／c小鼠体内毒力下降。仅SL7207在体外可向RAW264．7巨噬细胞递送真核表达质粒。SL7207的构建为重组沙门氏菌疫苗载体的研制提供了一个新的选择。     

Comparison of sexual compatibility in crosses between the southern and northern populations of the cabbage beetle Colaphellus bowringi

It is widely accepted that the genetic divergence and reproductive incompat- ibility between closely related species and/or populations is often viewed as an important step toward speciation. In this study, sexual compatibility in crosses between the southern XS population and the northern TA population of the polyandrous cabbage beetle Co- laphellus bowringi was investigated by testing their mating preferences, mating latency, copulation duration, and reproductive performances of post-mating. In choice mating ex- periments, the percentages ofmatings were significantly higher in intra-population crosses than in inter-population crosses. Both isolation index （/） and index of pair sexual isolation （/PSi） indicated partial mating incompatibility or assortative mating in crosses between the two different geographical populations. In single pair mating experiments, XS females in inter-population crosses mated significantly later and copulated significantly shorter than those in intra-population crosses. However, TA females in inter-population crosses mated significantly earlier and copulated longer than those in intra-population crosses, suggesting that larger XS males may enhance heterotypic mating. The lifetime fecundity was highest in XS homotypic matings, lowest in TA homotypic matings, and intermedi- ate in heterotypic rnatings between their parents. The inter-population crosses resulted in significantly lower egg hatching rate and shorter female longevity than intra-population crosses. These results demonstrated that there exist some incompatibilities in premating, postmating-prezygotic, and postzygotic stages between the southern XS population and northern TA population of the cabbage beetle Colaphellus bowringi.     

The Association between Genetic Polymorphism and the Processing Efficiency of miR-149 Affects the Prognosis of Patients with Head and Neck Squamous Cell Carcinoma

MicroRNAs (miRNAs) play important roles in modulating the neoplastic process of cancers including head and neck squamous cell carcinoma (HNSCC). A genetic polymorphism (rs2292832, C>T) has been recently identified in the precursor of miR-149; nevertheless its clinicopathological implications remain obscure. In this study, we showed that miR-149 is down-regulated in HNSCC compared to normal mucosa and this is associated with a poorer patient survival. In addition, HNSCC patients with the T/T genotype have more advanced tumors and a worse prognosis. Multivariate analysis indicated that patients carried the T/T genotype have a 2.81-fold (95% CI: 1.58–4.97) increased risk of nodal metastasis and 1.66-fold (95% CI: 1.05–2.60) increased risk of mortality compared to other groups. T/T genotype also predicted the worse prognosis of buccal mucosa carcinoma subset of HNSCC. In vitro analysis indicated that exogenous miR-149 expression reduces the migration of HNSCC cells. Moreover, HNSCC cell subclones carrying the pri-mir-149 sequence containing the T variant show a low processing efficacy when converting the pre-mir-149 to mature miR-149. These findings suggest that miR-149 suppresses tumor cell mobility, and that the pre-mir-149 polymorphism may affect the processing of miR-149, resulting in a change in the abundance of the mature form miRNA, which, in turn, modulates tumor progression and patient survival.     

Adoptive transfers of CD4+CD25+ Tregs partially alleviate mouse premature ovarian insufficiency

This study was designed to investigate the protective effect of CD4⁺CD25⁺ regulatory T cells (Tregs) against zona pellucida glycoprotein 3 peptide (pZP3) immunization‐induced premature ovarian insufficiency (POI) in mice. A mouse POI model was induced by two subcutaneous injections of pZP3 (50 nmol/L). Mice in the pZP3‐Treg group were intraperitoneally injected with 5 × 10⁵ CD4⁺CD25⁺ Tregs after the POI model was established. Sex hormone levels, follicle numbers, apoptotic events, and the Akt/FOXO3a signaling pathway molecules in the ovaries were assessed. Compared with control group, the weight of ovaries in both pZP3 group and pZP3‐Treg group was decreased and no difference was found between them. The number of follicles in the Treg transferred mice, like in pZP3 group, was significantly reduced compared to the control group, but showed a modest improvement when compared the pZP3 group alone. Significantly lower serum concentrations of follicle‐stimulating hormone, luteinizing hormone, and anti‐zona pellucida antibodies (AZPAbs) were found, while the concentrations of estradiol and anti‐Mullerian hormone increased. In mechanism, Treg cell transfer to ZP3 treated mice restored the levels of Caspase3 to control levels, and partially restored Bax, however, had no effect on Bcl‐2. Moreover, Treg cell transfer to ZP3 treated mice partially restored the levels of Akt and FOXO3a, and partially restored the ratios of p‐Akt/Akt and p‐FOXO3a/FOXO3a. In conclusion, Treg cells improved some aspects of ZP3‐induced POI which may be mediate by suppressing ovarian cells apoptosis and involving the Akt/FOXO3a signaling pathway. Therefore, Treg cells may be protective against autoimmune POI.     

The crosstalk between lncRNAs and the Hippo signalling pathway in cancer progression

LncRNAs play a pivotal role in the regulation of epigenetic modification, cell cycle, differentiation, proliferation, migration and other physiological activities. In particular, considerable studies have shown that the aberrant expression and dysregulation of lncRNAs are widely implicated in cancer initiation and progression by acting as tumour promoters or suppressors. Hippo signalling pathway has attracted researchers’ attention as one of the critical cancer‐related pathways in recent years. Increasing evidences have demonstrated that lncRNAs could interact with Hippo cascade and thereby contribute to acquisition of multiple malignant hallmarks, including proliferation, metastasis, relapse and resistance to anti‐cancer treatment. Specifically, Hippo signalling pathway is reported to modulate or be regulated by widespread lncRNAs. Intriguingly, certain lncRNAs could form a reciprocal feedback loop with Hippo signalling. More speculatively, lncRNAs related to Hippo pathway have been poised to become important putative biomarkers and therapeutic targets in human cancers. Herein, this review focuses on the crosstalk between lncRNAs and Hippo pathway in carcinogenesis, summarizes the comprehensive role of Hippo‐related lncRNAs in tumour progression and depicts their clinical diagnostic, prognostic or therapeutic potentials in tumours.     

Feedback regulation of coronary artery disease susceptibility gene ADTRP and LDL receptors LDLR/CD36/LOX-1 in endothelia cell functions involved in atherosclerosis

A high level of low-density lipoprotein cholesterol (LDL) is one of the most important risk factors for coronary artery disease (CAD), the leading cause of death worldwide. However, a low concentration of LDL may be protective. Genome-wide association studies revealed that variation in ADTRP gene increased the risk of CAD. In this study, we found that a low concentration of oxidized-LDL induced the expression of ADTRP. Further analyses showed that knockdown of the expression of LDL receptor genes LDLR, CD36, or LOX-1 significantly downregulated ADTRP expression, whereas overexpression of LDLR/CD36/LOX-1 markedly increased ADTRP expression through the NF-κB pathway. Like ADTRP, LDLR, CD36 and LOX-1 were all involved in endothelial cell (EC) functions relevant to the initiation of atherosclerosis. Downregulation of LDLR/CD36/LOX-1 promoted monocyte adhesion to ECs and transendothelial migration of monocytes by increasing expression of ICAM-1, VCAM-1, E-selectin and P-selectin, decreased EC proliferation and migration, and increased EC apoptosis, thereby promoting the initiation of atherosclerosis. Opposite effects were observed with the overexpression of ADTRP and LDLR/CD36/LOX-1 in ECs. Interestingly, through the NF-κB and AKT pathways, overexpression of ADTRP significantly upregulated the expression of LDLR, CD36, and LOX-1, and knockdown of ADTRP expression significantly downregulated the expression of LDLR, CD36, and LOX-1. These data suggest that ADTRP and LDL receptors LDLR/CD36/LOX-1 positively regulate each other, and form a positive regulatory loop that regulates endothelial cell functions, thereby providing a potential protective mechanism against atherosclerosis. Our findings provide a new molecular mechanism by which deregulation of ADTRP and LDLR/CD36/LOX-1 promote the development of atherosclerosis and CAD.     

Standardized xeno- and serum-free culture platform enables large-scale expansion of high-quality mesenchymal stem/stromal cells from perinatal and adult tissue sources

Background aimsMesenchymal stem/stromal cells (MSCs) are of interest for the treatment of graft-versus-host disease, autoimmune diseases, osteoarthritis and neurological and cardiovascular diseases. Increasing numbers of clinical trials emphasize the need for standardized manufacturing of these cells. However, many challenges related to diverse isolation and expansion protocols and differences in cell tissue sources exist. As a result, the cell products used in numerous trials vary greatly in characteristics and potency.MethodsThe authors have established a standardized culture platform using xeno- and serum-free commercial media for expansion of MSCs derived from umbilical cord (UC), bone marrow and adipose-derived (AD) and examined their functional characteristics.ResultsMSCs from the tested sources stably expanded in vitro and retained their biomarker expression and normal karyotype at early and later passages and after cryopreservation. MSCs were capable of colony formation and successfully differentiated into osteogenic, adipogenic and chondrogenic lineages. Pilot expansion of UC-MSCs and AD-MSCs to clinical scale revealed that the cells met the required quality standard for therapeutic applications.ConclusionsThe authors’ data suggest that xeno- and serum-free culture conditions are suitable for large-scale expansion and enable comparative study of MSCs of different origins. This is of importance for therapeutic purposes, especially because of the numerous variations in pre-clinical and clinical protocols for MSC-based products.