A Review on Investigating the Interactions between Nanoparticles and the Pulmonary Surfactant Monolayer with Coarse-Grained Molecular Dynamics Method.

Pulmonary drug delivery has garnered significant attention due to its targeted local lung action, minimal toxic side effects, and high drug utilization. However, the physicochemical properties of inhaled nanoparticles (NPs) used as drug carriers can influence their interactions with the pulmonary surfactant (PS) monolayer, potentially altering the fate of the NPs and impairing the biophysical function of the PS monolayer. Thus, the objective of this review is to summarize how the physicochemical properties of NPs affect their interactions with the PS monolayer. Initially, the definition and properties of NPs, as well as the composition and characteristics of the PS monolayer, are introduced. Subsequently, the coarse-grained molecular dynamics (CGMD) simulation method for studying the interactions between NPs and the PS monolayer is presented. Finally, the implications of the hydrophobicity, size, shape, surface charge, surface modification, and aggregation of NPs on their interactions with the PS monolayer and on the composition of biomolecular corona are discussed. In conclusion, gaining a deeper understanding of the effects of the physicochemical properties of NPs on their interactions with the PS monolayer will contribute to the development of safer and more effective nanomedicines for pulmonary drug delivery.

Numerical investigations of translocation characteristics of nano-silica lunar dust across pulmonary surfactant monolayer.

The interactions between nano-silica lunar dust (NSLD) on the moon surface and pulmonary surfactant (PS) monolayer will pose risks to astronaut health in future manned lunar exploration missions, but the specifics of these interactions are unknown. This study investigates them using the coarse-grained molecular dynamics method considering different sizes (5, 10, and 15 nm) and shapes (sphere, ellipsoid, and cube), with special focus on the unique morphology of NSLDs with bugles. The key findings are as follows: (1) The 10 nm and 15 nm NSLDs embed in the PS monolayer through the major sphere of spherical-type, major ellipsoid of ellipsoidal-type, or one edge of cubic-type NSLDs upon contact the PS monolayer. (2) Adsorbed NSLDs cause a higher Sz value (ASz > 0.84), while embedded NSLDs cause a lower Sz value (0.47 < ASz < 0.83) that decreases with an increase in the number of bulges. (3) The embedding process absorbs 50-342 dipalmitoylphosphatidylcholine (DPPC) molecules, reducing the PS monolayer area by 0.21%-6.05%. NSLDs with bulges absorb approximately 9-126 additional DPPC molecules and cause a 0.05%-3.22% reduction in the PS monolayer area compared to NSLDs without bulges. (4) NSLDs move obliquely or vertically within the PS monolayer, displaying two distinct stages with varying velocities. Their movement direction and speed are influenced by the increasing complexity of NSLD with more bulges on them. In general, larger NSLDs with sharper shapes and increasing complex morphology of more bulges cause more significant damages to the PS monolayer. These findings have implications for safeguarding astronaut health in future manned lunar exploration missions.

SEMA3A Exon 9 Expression Is a Potential Prognostic Marker of Unfavorable Recurrence-Free Survival in Patients with Tongue Squamous Cell Carcinoma.

This study tried to assess the prognostic value of semaphorin (SEMA) family genes in patients with tongue squamous cell carcinoma (TSCC) and the potential epigenetic alterations of the genes. The part of third-level TSCC data in The Cancer Genome Atlas-Head and Neck Squamous Cell Carcinoma (TCGA-HNSC) was extracted using the UCSC Xena browser for analysis. Among 20 SEMA genes examined, 7 were markedly upregulated, while 8 were substantially decreased in TSCC tissues compared with adjacent normal tissues. SEMA3A was the only gene with independent prognostic value in terms of recurrence-free survival (RFS) in multivariate analysis (hazard ratio [HR]: 1.697, 95% CI: 1.228-2.345, p = 0.001). Among the individual exons of SEMA3A, the exon 9 had a better prognostic value in terms of recurrence than total SEMA3A expression and its expression also independently predicted shorter RFS (HR: 2.193, 95% CI: 1.463-3.290, p < 0.001). The methylation levels of two CpG sites (cg06144675 and cg13988052) were moderately correlated with SEMA3A expression. Interestingly, cg06144675, which locates at the promoter region, showed a negative correlation with SEMA3A expression, whereas cg13988052, which is in the intron of SEMA3A gene body showed a positive correlation with SEMA3A expression. In conclusion, SEMA3A expression is aberrantly upregulated in TSCC tissues. Its exon 9 expression is a potentially valuable prognostic marker of unfavorable RFS in TSCC patients. Both promoter hypomethylation and gene body hypermethylation might contribute to the dysregulation.

Substance P participates in periodontitis by upregulating HIF-1α and RANKL/OPG ratio.

BACKGROUND: Both substance P and hypoxia-inducible factor 1 alpha (HIF-1α) are involved in inflammation and angiogenesis. However, the relationship between substance P and HIF-1α in rat periodontitis is still unknown. METHODS: Ligation-induced rat periodontitis was established to observe the distribution and expression of substance P and HIF-1α by immunohistochemistry. Rat gingival fibroblasts were cultured and stimulated with Porphyromonas gingivalis lipopolysaccharide (LPS). Recombinant substance P was applied to elaborate the relationship between substance P and HIF-1α in gingival fibroblasts in vitro. Primary mouse bone marrow-derived macrophages (BMMs) were isolated and cultured to observe the effect of substance P on receptor activator of NF-κB ligand (RANKL)-induced osteoclastogenesis by TRAP staining. Western blotting was used to investigate the expression of HIF-1α, osteoprotegerin (OPG) and RANKL. RESULTS: Rat experimental periodontitis was successfully established 6 weeks after ligation. Gingival inflammatory infiltration and alveolar bone loss were observed. Positive expression of substance P was found in the infiltrating cells. Higher HIF-1α levels were observed in periodontitis compared to that of normal tissues. Substance P upregulated the level of HIF-1α in gingival fibroblasts with or without 1 µg/ml LPS in vitro (*P < 0.05). Substance P upregulated the expression of HIF-1α in RANKL-stimulated BMMs in vitro. Substance P also increased the RANKL/OPG ratio in gingival fibroblasts (*P < 0.05). Both 10 nM and 50 nM substance P promoted RANKL-induced osteoclast differentiation (*P < 0.05). CONCLUSION: Substance P participates in periodontitis by upregulating HIF-1α and the RANKL/OPG ratio.

MiR-638 suppresses the progression of oral squamous cell carcinoma through wnt/ß-catenin pathway by targeting phospholipase D1.

Objective: The current study aimed to explore the function of miR-638 on the progression of oral squamous cell carcinoma (OSCC) and relevant molecular mechanisms. Methods: Expression profile of miR-638 in OSCC tissues and cells was detected using quantitative real-time polymerase chain reaction (qRT-PCR) method. Chi-square test was performed to estimate the relationship between miR-638 and clinical parameters of OSCC cases. Cell viability and motility abilities were estimated using MTT and transwell assays, respectively. Potential targets of miR-638 in OSCC were identified through bioinformatics analysis and luciferase reporter assay. Results: MiR-638 exhibited decreased expression in OSCC tissues and cells, compared to non-cancerous controls (P < .05 for both). Moreover, its down-regulation was closely correlated with lymph node metastasis (P = .044) and TNM stages (P = .001). Enforced miR-638 expression reduced cell proliferation, migration and invasion, while its knockdown exhibited opposite effects. Phospholipase D1 (PLD1) was confirmed as a target of miR-638 in OSCC. MiR-638 could inhibit wnt/ß-catenin pathway through targeting PLD1, thus realizing its anti-tumour action in OSCC. Conclusion: MiR-638 may be a tumour suppressor in OSCC by targeting PLD1/Wnt/ß-catenin pathway.

Semaphorin 3A gets involved in the establishment of mouse tooth eruptive pathway.

The accurately establishment of the eruptive pathway is of vital importance. The mechanisms governing tooth eruption pathway remain little known. This study is to elucidate the roles of Semaphorin 3A (Sema 3A) in mouse tooth eruptive pathway. C57BL/6 mice (11-13 and 15-17 days after birth) were chosen to observe eruptive pathway of mouse lower first molar. Expressions of Sema 3A and its receptor neuropilin 1 and plexin A1 were detected. Osteoclasts were identified by TRAP staining. Co-localization of Sema 3A and osteoclast maker CD68 was detected by double immunofluorescence staining. Picrosirius red staining was applied to observe collagen fibers during mucosal penetration phase. In vitro, Bone marrow-derived macrophages (BMMs) were prepared from 4 week C57BL/6 mice to observe the effect of Sema 3A on the differentiation of BMMs into osteoclasts by TRAP staining. Expressions of Sema 3A was observed by immunofluorescence and western blotting. At osseous eruption phase, many TRAP-positive multi-nucleated cells were distributed around occlusal alveolar bone. The positive expressions of Sema 3A were observed in the multi-nucleated cells. Fluorescence double staining showed that Sema 3A and CD68 were co-expressed in osteoclasts. Its receptor neuropilin 1 and plexin A1 were also found in osteoclasts. In vitro, Sema3A negatively regulated osteoclast differentiation. At mucosal penetration, occlusal alveolar bone had been completely resorbed and collagen fires were gradually degraded for eruptive pathway. Similar positive expressions of Sema 3A and its receptor neuropilin 1 and plexin A1 were also found in the mucosal penetration pathway. Sema 3A gets involved in the establishment of mouse tooth eruptive pathway by modulating osteoclast activity. Sema3A should be considered as a novel nervous agent or a potential biomarker for mouse tooth eruptive pathway.

PAR-2 promotes cell proliferation, migration, and invasion through activating PI3K/AKT signaling pathway in oral squamous cell carcinoma.

Objective: This research aimed to explore the function of protease activated receptor 2 (PAR-2) in oral squamous cell carcinoma (OSCC) development and progression, as well as underlying molecular mechanism.Methods: Tissue samples were collected from 115 OSCC patients. Quantitative real-time PCR (qRT-PCR) was performed to measure the expression of PAR-2 mRNA in OSCC tissues and cells. MTT and Transwell assays were used to detect the proliferation, migration, and invasion of OSCC cells, respectively. Western blot was performed to determine protein expression.Results: The expression of PAR-2 mRNA was up-regulated in OSCC tissue and cells (P<0.01), and its mRNA level was obviously correlated to tumor differentiation and TNM stage in OSCC (P<0.05 for both). The activation of PAR-2 with PAR-2AP (PAR-2 agonist) significantly promoted the proliferation, migration, and invasion of OSCC cells, while its knockout could inhibit malignant behaviors of OSCC cells (P<0.05). Excessive activation of PAR-2 enhanced phosphorylation level of PI3K, AKT, and mTOR revealing the activation of PI3K/AKT pathway. Moreover, LY294002, the inhibitor of PI3K/AKT pathway, could reverse oncogenic action caused by PAR-2 activation.Conclusion:PAR-2 can promote OSCC growth and progression via activating PI3K/AKT signaling pathway.

The effects of Sema3A overexpression on the proliferation and differentiation of rat gingival mesenchymal stem cells in the LPS-induced inflammatory environment.

Semaphorin3A has been identified as a potent osteoprotective factor that simultaneously inhibits bone resorption and promotes bone formation. The present study demonstrates the effect of the overexpression of Sema3A on the proliferation and differentiation of rat gingival mesenchymal stem cells (GMSCs) in the lipopolysaccharide (LPS)-induced inflammatory environment. rGMSCs were transfected with viral stocks of pLenO-GTP-Sema3A (Lv-Sema3A group) or pLenO-GTP (Lv-NC group), with rGMSCs as a control. The transfection efficiency was determined by flow cytometry. Cell proliferation was assessed using a Cell Counting Kit-8 assay. The expressions of alkaline phosphatase (ALP), osteocalcin (OCN), and runt-related transcription factor 2 (Runx2) were determined at 3, 7 and 14 days after the osteogenic induction culture with or without LPS using real-time PCR and Western blot. Alizarin Red staining was performed at 28 days. A pLenO-GTP-Sema3A-mediated transfection of rGMSC stably overexpressing Sema3A was built up. The overexpression of Sema3A promoted cell proliferation in the LPS-induced inflammatory environment. In addition, osteogenesis-related genes were upregulated in the Lv-Sema3A group compared with the control group. Also, after LPS administration, the overexpression of Sema3A enhanced the expression of the osteogenic genes in the LPS-induced inflammatory environment. Hence, Sema3A gene-modified rGMSCs show better osteogenic differentiation and proliferation capacities compared with rGMSCs in the LPS-induced inflammatory environment.

[Effect of hypoxia on the expression of HIF-1alpha and VEGF in human periodontal ligament cells in vitro].

PURPOSE: To investigate the effect of hypoxia on the expression of HIF-1alpha and VEGF in human periodontal ligament cells (hPDLCs) in vitro. METHODS: HPDLCs were cultured in Dulbecco's modified Eagle's medium (DMEM) and subcultured at confluence. Experiments were carried out with the third passage of hPDLCs populations. In the normoxic control group, cells were incubated under normoxic conditions of 20% O2, 5% CO2, 75% N2 for 24 hours and 48 hours, respectively. In the hypoxic group, cells were incubated in a humidified atmosphere of 1% O2, 5% CO2, 94% N2 for 24 hours and 48 hours, respectively. The expression of HIF-1alpha and VEGF in hPDLCs in vitro was measured using reverse transcription-polymerase chain reaction. SP immunohistochemistry method was performed to localize the distribution of HIF-1alpha and VEGF in hPDLCs. The data was analyzed by Student's t test, one-way ANOVA and LSD test with SPSS 13.0 software package. RESULTS: There was no significant difference between the hypoxic group and the normoxic control group in the expression of HIF-1alpha mRNA in hPDLCs. The expression of VEGF mRNA in the hypoxic group of 24 hours and 48 hours was both statistically higher than in the control group (P<0.05). The expression of HIF-1alpha protein was significantly increased with hypoxia time while the staining of HIF-1alpha was negative under normoxic condition. The expression of VEGF protein in hypoxic groups was significantly higher than in normoxic control groups. The staining of VEGF gradually enhanced in a time-dependent manner and was weakly positive after 48 hours under normoxic condition. CONCLUSION: Hypoxia could change the metabolic pathway of human periodontal ligament cells by upregulating the expression of HIF-1alpha and other relevant growth factors.

[Effect of epidermal growth factor on transcription of MCP-1 in rat dental follicle cells].

PURPOSE: To study the effects of epidermal growth factor (EGF) on the secretion of monocyte chemotactic protein-one (MCP-1) in rat dental follicle cells. METHODS: Dental follicle was harvested from 4-6 days postnatal Wistar rats and cultured in alpha-MEM with 15% FBS. Cells in the third passage were incubated with EGF at different concentrations to choose the best dosage. MTT assays were used to evaluate cell proliferation. Using the suitable concentration, 10 ng/ml EGF cells were cultured for 0, 0.5, 1, 3 and 6 hours respectively, and then collected for RT-PCR analysis. The results were analyzed with SPSS13.0 software package. RESULTS: EGF(ng/ml) at 5-10 ng/ml increased the cell proliferation (P<0.05), which reached the peak at 10 ng/ml according to MTT assay (P<0.01). 10 ng/ml and 3 hours EGF exposure up-regulated the cellular MCP-1 expression dramatically (P<0.01). CONCLUSION: Proper concentration of EGF may promote dental follicle cell proliferation, EGF may up-regulate MCP-1 transcription in the rat dental follicle cells.

[Proliferation, apoptosis and expression of tissue non-specific alkaline phosphatase in cells of stratum intermedium during amelogenesis in postnatal stage of BALB/c mouse].

PURPOSE: To study the apoptosis and expression of proliferating cell nuclear antigen(PCNA),tissue non-specific alkaline phosphatase(TNSALP) of stratum intermedium cell in postnatal stage of BALB/c mouse. METHODS: Fifty six BALB/c mice in different developmental days were sacrificed and their bilateral mandibular first molar germs with surrounding alveolar bone were taken out, then the tissues were fixed with 4% paraformaldehyde at 4degrees centigrade overnight, dehydrated, embedded in paraffin and serially sectioned at 5mum.Tdt-mediated Dutp nick end labeling (TUNEL) and immunohistochemical assay were adopted to determine the tissue distribution and cellular localization of bcl-2, PCNA and TNSALP,respectively. Image-pro plus 6.0 software was used to evaluate the histological sections and the data was analyzed by Students't test,one-way ANOVA and SNK-q test with SPSS 13.0 software package. RESULTS: The expression of PCNA in stratum intermedium was higher than ameloblast at postnatal day 1, but it gradually decreased and was negative at postnatal day 7. The apoptosis of stratum intermedium cells increased during enamel development.The expression of TNSALP appeared at postnatal day 3 and was most obvious at postnatal day 5 in stratum intermedium, then gradually decreased. However, the expression of TNSALP appeared at postnatal day 7 in ameloblasts, then gradually increased. CONCLUSIONS: There is a significant linear correlation between stratum intermedium cell and ameloblast in apoptosis and expression of PCNA and TNSALP.It is suggested that the stratum intermedium participate in the differentiation of ameloblast and formation of enamel.