Complete genome sequence of Schaalia odontolytica isolated from subgingival biofilm.

OBJECTIVE: Recent advancements in genome-based taxonomic classification propose the reclassification of certain Actinomyces species into new genera, including Schaalia. Schaalia odontolytica, the type species within this genus, is frequently found in the human oral cavity and has been associated with actinomycotic lesions. Currently, only two complete genomes of S. odontolytica strains have been reported. Recognizing the limited research on subspecies-level variation of S. odontolytica, we conducted genome sequencing of strain KHUD_008, isolated from a Korean periodontitis patient's subgingival biofilm. Additionally, we performed a comparative genome analysis using previously sequenced genomes of strain XH001 and strain FDAARGOS_732, both derived from the human oral cavity. DATA DESCRIPTION: Pacific Biosciences Sequel II sequencing generated 15,904 and 76,557 raw sequencing sub-reads, which were integrated to assemble the de novo genome using the Microbial Genome Analysis pipeline in the Single-Molecule Real-Time Analysis. The genome assembly completeness, assessed by Benchmarking Universal Single-Copy Orthologs, reached 99.2%. The genome is 2,389,595 bp with a GC content of 66.37%, and contains 2,002 protein-coding genes, 9 rRNAs, and 48 tRNA. Comparative analysis with two previously sequenced strains revealed many strain-specific genes in KHUD_008, primarily related to envelope biogenesis and replication/recombination/repair processes.

Draft genome sequence of Limosilactobacillus fermentum isolated from subgingival biofilm.

Limosilactobacillus fermentum is generally considered beneficial for vaginal and digestive health. However, strains isolated from the oral cavity, especially from periodontitis lesions, have not been thoroughly studied. Here, we report the draft genome sequence of strain KHUD_007 isolated from the subgingival biofilm of a Korean patient with periodontitis.

Single-cell RNA sequencing reveals heterogeneity of adipose tissue-derived mesenchymal stem cells under chondrogenic induction.

This study investigated how adipose tissue-derived mesenchymal stem cells (AT-MSCs) respond to chondrogenic induction using droplet-based single-cell RNA sequencing (scRNA-seq). We analyzed 37,219 high-quality transcripts from control cells and cells induced for 1 week (1W) and 2 weeks (2W). Four distinct cell clusters (0-3), undetectable by bulk analysis, exhibited varying proportions. Cluster 1 dominated in control and 1W cells, whereas cluster 3, 2, and 0 exclusively dominated in control, 1W, and 2W cells, respectively. Furthermore, heterogeneous chondrogenic markers expression within clusters emerged. Gene ontology (GO) enrichment analysis of differentially expressed genes unveiled cluster-specific variations in key biological processes (BP): (1) Cluster 1 exhibited upregulation of GO-BP terms related to ribosome biogenesis and translational control, crucial for maintaining stem cell properties and homeostasis; (2) Additionally, cluster 1 showed upregulation of GO-BP terms associated with mitochondrial oxidative metabolism; (3) Cluster 3 displayed upregulation of GO-BP terms related to cell proliferation; (4) Clusters 0 and 2 demonstrated similar upregulation of GO-BP terms linked to collagen fibril organization and supramolecular fiber organization. However, only cluster 0 showed a significant decrease in GO-BP terms related to ribosome production, implying a potential correlation between ribosome regulation and the differentiation stages of AT-MSCs. Overall, our findings highlight heterogeneous cell clusters with varying balances between proliferation and differentiation before and after chondrogenic stimulation. This provides enhanced insights into the single-cell dynamics of AT-MCSs during chondrogenic differentiation.

Geranylgeraniol Application in Human Osteoblasts and Osteoclasts for Reversal of the Effect of Bisphosphonates.

Nitrogen-containing bisphosphonates lead to the depletion of geranylgeranyl pyrophosphate involved in the mevalonate pathway. The effect of geranylgeraniol (GGOH) on human osteoblast and osteoclast activities suppressed by zoledronate was investigated in this study. The effect of GGOH on human osteoblasts and osteoclasts subjected to treatment with zoledronate was analyzed by assessing cell viability, osteoclast differentiation, resorption ability, gene expression, and protein synthesis. Cell viability suppressed by bisphosphonates in osteoblasts and osteoprogenitor cells was restored with GGOH. Osteoclast differentiation was analyzed by vitronectin receptor immunofluorescence staining, and the addition of GGOH to zoledronate significantly increased osteoclast differentiation compared with zoledronate alone. A trend of reversal of osteoclast resorption by GGOH was observed; however, it was not significant in all groups. The expression of ALP, type 1 collagen, and RUNX2 in osteoblasts was recovered by the addition of GGOH. Only CALCR expression in osteoclasts was significantly recovered by GGOH addition in the zoledronate group. Although the activities of osteoblasts and osteoclasts were not entirely restored, the possibility that the topical application of GGOH in MRONJ patients or patients with dental problems and bisphosphonates might lessen the risk of development and recurrence of MRONJ is shown.

Comparative Transcriptome Analysis of Human Adipose-Derived Stem Cells Undergoing Osteogenesis in 2D and 3D Culture Conditions.

Human adipose-derived stem cells (hADSCs) are types of mesenchymal stem cells (MSCs) that have been used as tissue engineering models for bone, cartilage, muscle, marrow stroma, tendon, fat and other connective tissues. Tissue regeneration materials composed of hADSCs have the potential to play an important role in reconstituting damaged tissue or diseased mesenchymal tissue. In this study, we assessed and investigated the osteogenesis of hADSCs in both two-dimensional (2D) and three-dimensional (3D) culture conditions. We confirmed that the hADSCs successfully differentiated into bone tissues by ARS staining and quantitative RT-PCR. To gain insight into the detailed biological difference between the two culture conditions, we profiled the overall gene expression by analyzing the whole transcriptome sequencing data using various bioinformatic methods. We profiled the overall gene expression through RNA-Seq and further analyzed this using various bioinformatic methods. During differential gene expression testing, significant differences in the gene expressions between hADSCs cultured in 2D and 3D conditions were observed. The genes related to skeletal development, bone development and bone remodeling processes were overexpressed in the 3D culture condition as compared to the 2D culture condition. In summary, our RNA-Seq-based study proves effective in providing new insights that contribute toward achieving a genome-wide understanding of gene regulation in mesenchymal stem cell osteogenic differentiation and bone tissue regeneration within the 3D culture system.

Osteogenic differentiation and inflammatory response of recombinant human bone morphogenetic protein-2 in human maxillary sinus membrane-derived cells.

The aim of the present study was to investigate the osteogenic potential of human maxillary sinus membrane (hMSM)-derived cells, and the role of recombinant human bone morphogenetic protein-2 (rhBMP-2) in the inflammatory response of hMSM-derived cells and gingival fibroblasts following sinus floor elevation procedure (SFE). hMSM-derived cells from the samples were isolated, subcultured, and analyzed using immunohistochemical staining and flow cytometry. The hMSM-derived cells obtained from passage 6 were used for Alizarin Red staining and quantitative reverse transcription-quantitative PCR to observe its osteogenic activity and inflammatory reaction upon supplementation with rhBMP-2. The hMSM-derived cells were shown to be heterogeneous, as indicated by their positive expression of human mesenchymal stem cell markers (STRO-1, high mobility group AT-hook 2, CD44, CD105 and OCT-3/4), fibroblast cell marker (fibroblast-specific protein 1) and epithelial cell marker (epithelial cell adhesion molecule). Calcium nodules were found to be more notably evident in the rhBMP-2 group, following osteogenic differentiation. The gene expression of osteogenic markers was significantly upregulated in the cells supplemented with rhBMP-2. Supplementation with rhBMP-2 also enhanced the expression of inflammatory markers in hMSM-derived cells and gingival fibroblasts; however, NF-κB and TNF-α expression was not significantly increased compared with the control in the hMSM-derived cells. hMSM contains mesenchymal stem cells (MSCs) capable of differentiating into osteogenic cells. The supplementation of rhBMP-2 enhanced osteogenic differentiation and induced an inflammatory response which was greater in gingival fibroblasts compared with hMSM-derived cells. In summary, the hMSM is a potential contributor to the osteogenic process following SFE, and the use of rhBMP-2 may increase the inflammatory response accordingly. The gingival tissue may be responsible for the increased inflammatory response by rhBMP-2 and postoperative complications.

Identification of a novel Shank2 transcriptional variant in Shank2 knockout mouse model of autism spectrum disorder.

Autism spectrum disorder (ASD) is a group of neurodevelopmental disorders that are highly heterogeneous in clinical symptoms as well as etiologies. Mutations in SHANK2 are associated with ASD and accordingly, Shank2 knockout mouse shows ASD-like behavioral phenotypes, including social deficits. Intriguingly, two lines of Shank2 knockout (KO) mouse generated by deleting different exons (exon 6-7 or exon 7) showed distinct cellular phenotypes. Previously, we compared gene expressions between Shank2 KOs lacking exon 6-7 (e6-7 KO) and KOs lacking exon 7 (e7 KO) by performing RNA-seq. In this study, we expanded transcriptomic analyses to identify novel transcriptional variants in the KO mice. We found prominent expression of a novel exon (exon 4' or e4') between the existing exons 4 and 5 in the Shank2 e6-7 KO model. Expression of the transcriptional variant harboring this novel exon was confirmed by RT-PCR and western blotting. These findings suggest that the novel variant may function as a modifier gene, which contributes to the differences between the two Shank2 mutant lines. Furthermore, our result further represents an example of genetic compensation that may lead to phenotypic heterogeneity among ASD patients with mutations in the same gene.