低强度超声对鼻咽癌CNE2细胞增值和凋亡的影响

目的：观察低强度超声对人鼻咽癌CNF2细胞增殖和凋亡的影响。方法：以低强度超声（频率1．7MHz,剂量1．35W／cm2）辐照CNF2细胞,采用MTT观察超声对COCl细胞增殖的抑制作用;光镜观察细胞形态,细胞核荧光染色法检测凋亡细胞。结果：超声波能够显著抑制鼻咽癌CNE2细胞增殖,辐照后孵育18h,其MTT光吸收值与对照组相比监著降低：光镜下可见细胞形态学改变,同时Hoechst 33258荧光染色法观察到亮蓝色着染的凋亡细胞。结论：低强度超声辐照对人鼻咽癌CNE2细胞体外生长具有明显的抑制作用,并可诱导其凋亡。     

Octanoic Acid—An Insecticidal Metabolite of Conidiobolus coronatus (Entomopthorales) That Affects Two Majors Antifungal Protection Systems in Galleria mellonella (Lepidoptera): Cuticular Lipids and Hemocytes

The food flavour additive octanoic acid (C8:0) is also a metabolite of the entomopathogenic fungus Conidiobolus coronatus, which efficiently infects and rapidly kills Galleria mellonella. GC-MS analysis confirmed the presence of C8:0 in insecticidal fraction FR3 extracted from C. coronatus filtrate. Topical administration of C8:0 had a dose-dependent effect on survival rates of larvae but not on pupation or adult eclosion times of the survivors. Topically applied C8:0 was more toxic to adults than larvae (LD100 for adults 18.33 ± 2.49 vs. 33.56 ± 2.57 µg/mg of body mass for larvae). The administration of C8:0 on the cuticle of larvae and adults, in amounts corresponding to their LD50 and LD100 doses, had a considerable impact on the two main defense systems engaged in protecting against pathogens, causing serious changes in the developmental-stage-specific profiles of free fatty acids (FFAs) covering the cuticle of larvae and adults and damaging larval hemocytes. In vitro cultures of G. mellonella hemocytes, either directly treated with C8:0 or taken from C8:0 treated larvae, revealed deformation of hemocytes, disordered networking, late apoptosis, and necrosis, as well as caspase 1–9 activation and elevation of 8-OHdG level. C8:0 was also confirmed to have a cytotoxic effect on the SF-9 insect cell line, as determined by WST-1 and LDH tests.     

Revealing the Immune Heterogeneity between Systemic Lupus Erythematosus and Rheumatoid Arthritis Based on Multi-Omics Data Analysis

The pathogenesis of systemic lupus erythematosus (SLE) and rheumatoid arthritis (RA) are greatly influenced by different immune cells. Nowadays both T-cell receptor (TCR) and B-cell receptor (BCR) sequencing technology have emerged with the maturity of NGS technology. However, both SLE and RA peripheral blood TCR or BCR repertoire sequencing remains lacking because repertoire sequencing is an expensive assay and consumes valuable tissue samples. This study used computational methods TRUST4 to construct TCR repertoire and BCR repertoire from bulk RNA-seq data of both SLE and RA patients’ peripheral blood and analyzed the clonality and diversity of the immune repertoire between the two diseases. Although the functions of immune cells have been studied, the mechanism is still complicated. Differentially expressed genes in each immune cell type and cell–cell interactions between immune cell clusters have not been covered. In this work, we clustered eight immune cell subsets from original scRNA-seq data and disentangled the characteristic alterations of cell subset proportion under both SLE and RA conditions. The cell–cell communication analysis tool CellChat was also utilized to analyze the influence of MIF family and GALECTIN family cytokines, which were reported to regulate SLE and RA, respectively. Our findings correspond to previous findings that MIF increases in the serum of SLE patients. This work proved that the presence of LGALS9, PTPRC and CD44 in platelets could serve as a clinical indicator of rheumatoid arthritis. Our findings comprehensively illustrate dynamic alterations in immune cells during pathogenesis of SLE and RA. This work identified specific V genes and J genes in TCR and BCR that could be used to expand our understanding of SLE and RA. These findings provide a new insight inti the diagnosis and treatment of the two autoimmune diseases.     

Morphological and Biological Evaluations of Human Periodontal Ligament Fibroblasts in Contact with Different Bovine Bone Grafts Treated with Low-Temperature Deproteinisation Protocol

Several types of deproteinised bovine bone mineral (DBBM) are available on the market, and each one is obtained with a thermic and chemical process that can differ, achieving different results. Currently, several protocols using low temperature are suggested to reduce the possible particle crystallisation during the production process. This study aimed to evaluate the biomorphological reaction of periodontal fibroblast cultures in contact with different DBBM particles treated with a low-temperature protocol (Thermagen^®) and without exposure to sodium hydroxide (NaOH). Morphological evaluation was performed using light, confocal laser, and scanning electron microscopy, and the biological reaction in terms of proliferation was performed using an XTT proliferation assay at 24 h (T1), 72 h (T2), and 7 days (T3). The morphological analysis highlighted how the presence of the materials stimulated a change in the morphology of the cells into a polygonal shape, surface reactions with the thickening of the membrane, and expression of actin. In particular, the morphological changes were appreciable from T1, with a progressive increase in the considered morphological characteristics at T2 and T3 follow-ups. The proliferation assay showed a statistical significance between the different experimental materials and the negative control in T2 and T3 follow-ups. The post hoc analysis did not reveal any differences between the materials. In conclusion, the grafts obtained with the low-temperature extractions protocol and not exposed to NaOH solution showed positive morphological reactions with no differences in the sizes of particles.     

HIV-2 Neutralization Sensitivity in Relation to Co-Receptor Entry Pathways and Env Motifs

HIV-2, compared to HIV-1, elicits potent and broadly neutralizing antibodies, and uses a broad range of co-receptors. However, both sensitivity to neutralization and breadth of co-receptor use varies between HIV-2 isolates, and the molecular background is still not fully understood. Thus, in the current study, we have deciphered relationships between HIV-2 neutralization sensitivity, co-receptor use and viral envelope glycoprotein (Env) molecular motifs. A panel of primary HIV-2 isolates, with predefined use of co-receptors, was assessed for neutralization sensitivity using a set of HIV-2 Env-directed monoclonal antibodies and co-receptor indicator cell lines. Neutralization sensitivity of the isolates was analysed in relation target cell co-receptor expression, in addition to amino acid motifs and predicted structures of Env regions. Results showed that HIV-2 isolates were more resistant to neutralizing antibodies when entering target cells via the alternative co-receptor GPR15, as compared to CCR5. A similar pattern was noted for isolates using the alternative co-receptor CXCR6. Sensitivity to neutralizing antibodies appeared also to be linked to specific Env motifs in V1/V2 and C3 regions. Our findings suggest that HIV-2 sensitivity to neutralization depends both on which co-receptor is used for cell entry and on specific Env motifs. This study highlights the multifactorial mechanisms behind HIV-2 neutralization sensitivity.     

Sulfated Hyaluronan Binds to Heparanase and Blocks Its Enzymatic and Cellular Actions in Carcinoma Cells

We examined whether sulfated hyaluronan exerts inhibitory effects on enzymatic and biological actions of heparanase, a sole endo-beta-glucuronidase implicated in cancer malignancy and inflammation. Degradation of heparan sulfate by human and mouse heparanase was inhibited by sulfated hyaluronan. In particular, high-sulfated hyaluronan modified with approximately 2.5 sulfate groups per disaccharide unit effectively inhibited the enzymatic activity at a lower concentration than heparin. Human and mouse heparanase bound to immobilized sulfated hyaluronan. Invasion of heparanase-positive colon-26 cells and 4T1 cells under 3D culture conditions was significantly suppressed in the presence of high-sulfated hyaluronan. Heparanase-induced release of CCL2 from colon-26 cells was suppressed in the presence of sulfated hyaluronan via blocking of cell surface binding and subsequent intracellular NF-κB-dependent signaling. The inhibitory effect of sulfated hyaluronan is likely due to competitive binding to the heparanase molecule, which antagonizes the heparanase-substrate interaction. Fragment molecular orbital calculation revealed a strong binding of sulfated hyaluronan tetrasaccharide to the heparanase molecule based on electrostatic interactions, particularly characterized by interactions of (−1)- and (−2)-positioned sulfated sugar residues with basic amino acid residues composing the heparin-binding domain-1 of heparanase. These results propose a relevance for sulfated hyaluronan in the blocking of heparanase-mediated enzymatic and cellular actions.