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Periodontal Ehlers–Danlos syndrome arising from heterozygous pathogenic mutation in C1R and/or C1S genes is an autosomal-dominant disorder characterized by early-onset periodontitis. Due to the difficulties in obtaining and culturing the patient-derived gingival fibroblasts, we established a model system by introducing a heterozygous C1RR301P/WT mutation into human TERT-immortalized gingival fibroblasts (hGFBs) to investigate its specific effects on collagen metabolism and inflammatory responses. A heterozygous C1RR301P/WT mutation was introduced into hGFBs using engineered prime editing. The functional consequences of this mutation were assessed at cellular, molecular, and enzymatic levels using a variety of techniques, including cell growth analysis, collagen deposition quantification, immunocytochemistry, enzyme-linked immunosorbent assay, and quantitative real-time reverse transcription polymerase chain reaction. The C1RR301P/WT-mutated hGFBs (mhGFBs) exhibited normal morphology and growth rate compared to wild-type hGFBs. However, mhGFBs displayed upregulated procollagen α1(V), MMP-1, and IL-6 mRNA expression while simultaneously downregulating collagen deposition and C1r protein levels. A modest accumulation of unfolded collagens was observed in mhGFBs. The mhGFBs exhibited a heightened inflammatory response, with a more pronounced increase in MMP-1 and IL-6 mRNA expression compared to TNF-α/IL-1β-stimulated hGFBs. Unlike cytokine-stimulated hGFBs, cytokine-stimulated mhGFB did not increase C1R, C1S, procollagen α1(III), and procollagen α1(V) mRNA expression. Our results suggest that the C1RR301P/WT mutation specifically disrupts collagen metabolism and inflammatory pathways in hGFBs, highlighting the mutation’s role in these processes. While other cellular functions appear largely unaffected, these findings underscore the potential of targeting collagen metabolism and inflammation for therapeutic interventions in pEDS.

Signal regulatory protein alpha (SIRPα) is mainly expressed by cells of myeloid origin. This membrane glycoprotein is shown to be involved in regulation of different inflammatory conditions, such as colitis and arthritis. However, SIRPα has not been investigated in relationship to periodontitis, an inflammatory condition affecting the tooth supporting tissues. We aim to investigate if resident cells in the periodontium express SIRPα and whether a possible expression is affected by inflammatory conditions. Primary human keratinocytes, fibroblasts, periodontal ligament cells, and osteoblasts were cultured with or without the pro-inflammatory cytokines tumor necrosis factor alpha (TNF-α) or interleukin-1-beta (IL-1β). All different periodontal cell types showed a basal mRNA expression of SIRPα. Pro-inflammatory cytokines induced a 2-3-fold significant increase in SIRPα expression in both cultured human gingival fibroblasts and osteoblasts but neither in keratinocytes nor in periodontal ligament cells. Tissue sections from human gingival tissue biopsies were histochemically stained for SIRPα. Epithelial keratinocytes and gingival fibroblasts stained positive in sections from periodontally healthy as well as in sections from periodontitis. In periodontitis sections, infiltrating leukocytes stained positive for SIRPα. We highlight our finding that oral keratinocytes, gingival fibroblasts, and periodontal ligament cells do express SIRPα, as this has not been presented before. The fact that inflammatory stimulation of gingival fibroblasts increased the expression of SIRPα, while an increased expression by gingival fibroblasts in periodontitis tissue in situ could not be detected, is indeed contradictory.

Background. Complications of orthognathic surgery are quite rare, but they cause suffering in affected individuals.The range of complications is broad and includes both hard and soft tissue.

Case presentation: We here present a case of a fully healthy woman without signs of impaired healing capacity. Thepatient underwent bimaxillary orthognathic surgery and experienced multiple complications both peri- and postoperatively.During the post operative period, the patient also suffered from soft tissue complications after an orthopaedicinjury. Therefore, we referred the patient to her general practitioner for further medical investigation. Wealso present the result after restorative surgery and endodontic and prosthodontic treatment resulting in a successfulrehabilitation.

Conclusion: This case report clearly shows the need for a good collaboration between different odontologicaland medical fields to achieve a good and predictable result. In situations where normal healing processesdo not occur, in-depth analysis must be carried out.

Highlights: Orthognathic surgery affects soft and hard tissue which can result in adverse healing and complications.It is of great importance to follow up performed surgery to see late complications. Be restrictive with earlyre-operations when there are signs of necrosis. Always use a multidisciplinary approach when handling complicationsafter surgery.

Although three-dimensional (3D) co-culture of gingival keratinocytes and fibroblasts-populated collagen gel can mimic 3D structure of in vivo tissue, the uncontrolled contraction of collagen gel restricts its application in clinical and experimental practices. We here established a stable 3D gingival tissue equivalent (GTE) using hTERT-immortalized gingival fibroblasts (hGFBs)-populated collagen gel directly crosslinked with genipin/cytochalasin D and seeding hTERT-immortalized gingival keratinocytes (TIGKs) on the upper surface for a 2-week air–liquid interface co-culture. MTT assay was used to measure the cell viability of GTEs. GTE size was monitored following culture period, and the contraction was analyzed. Immunohistochemical assay was used to analyze GTE structure. qRT-PCR was conducted to examine the mRNA expression of keratinocyte-specific genes. Fifty µM genipin (G50) or combination (G + C) of G50 and 100 nM cytochalasin D significantly inhibited GTE contraction. Additionally, a higher cell viability appeared in GTEs crosslinked with G50 or G + C. GTEs crosslinked with genipin/cytochalasin D showed a distinct multilayered stratified epithelium that expressed keratinocyte-specific genes similar to native gingiva. Collagen directly crosslinked with G50 or G + C significantly reduced GTE contraction without damaging the epithelium. In summary, the TIGKs and hGFBs can successfully form organotypic multilayered cultures, which can be a valuable tool in the research regarding periodontal disease as well as oral mucosa disease. We conclude that genipin is a promising crosslinker with the ability to reduce collagen contraction while maintaining normal cell function in collagen-based oral tissue engineering.

We recently showed that adult male mice that lacked the C–C-chemokine receptor 3 (CCR3) exhibited disturbed bone remodeling, which resulted in a cortical bone phenotype of thin femoral cortical bone. However, it remains unknown whether this phenotype would be present during bone modeling, or it affects female mice. Here, we analyzed juvenile and adolescent CCR3-deficient mice to determine when bone modeling was affected in the absence of CCR3 signaling. To investigate whether the CCR3 bone phenotype was sex-related, we analyzed both young female and male mice, and adult females.

Micro-computed tomography (μCT) and histomorphometric analyses in adolescent CCR3-deficient male mice revealed reduced cortical bone volume and thickness, and an increase in periosteal mineralization. Interestingly, no skeletal phenotype was observed in adolescent or adult female CCR3-deficient mice. Among juvenile CCR3-deficient mice, neither males nor females showed a skeletal phenotype, which indicated that bone modeling was not affected by the CCR3 deficiency.

In summary, adolescent and adult male mice that lacked CCR3 receptors exhibited a cortical phenotype that was not present in female mice, probably due to an estrogen protective mechanism. Based on these and our previous results, we suggest that the importance of CCR3 in cortical bone turnover is related to sex hormones. Because only a few molecules are known to control cortical bone turnover, our novel finding that CCR3 regulated cortical bone thickness only in males suggested that CCR3 is a novel target for controlling cortical bone morphology in male individuals, and perhaps, in post-menopausal women.

Increasing evidence emphasizes the importance of chemokines and chemokine receptors as regulators of bone remodeling. The C–C chemokine receptor 3 (CCR3) is dramatically upregulated during osteoclastogenesis, but the role of CCR3 in osteoclast formation and bone remodeling in adult mice is unknown. Herein, we used bone marrow macrophages derived from adult male CCR3-proficient and CCR3-deficient mice to study the role of CCR3 in osteoclast formation and activity. CCR3 deficiency was associated with formation of giant hypernucleated osteoclasts, enhanced bone resorption when cultured on bone slices, and altered mRNA expression of related chemokine receptors and ligands. In addition, primary mouse calvarial osteoblasts isolated from CCR3-deficient mice showed increased mRNA expression of the osteoclast activator–related gene, receptor activator of nuclear factor kappa-B ligand, and osteoblast differentiation–associated genes. Microcomputed tomography analyses of femurs from CCR3-deficient mice revealed a bone phenotype that entailed less cortical thickness and volume. Consistent with our in vitro studies, the total number of osteoclasts did not differ between the genotypes in vivo. Moreover, an increased endocortical osteoid mineralization rate and higher trabecular and cortical bone formation rate was displayed in CCR3-deficient mice. Collectively, our data show that CCR3 deficiency influences osteoblast and osteoclast differentiation and that it is associated with thinner cortical bone in adult male mice.

BACKGROUND: Innate lymphoid cells (ILCs) are the most recently identified leukocytes of the immune system and these cells are increasingly acknowledged to play important roles in host defence and tissue repair. ILCs are also contributors of inflammatory diseases such as asthma and colitis. We analyzed the presence and relative proportions of the different ILC subsets (ILC1, ILC2 and ILC3) in gingivitis and periodontitis. Further, we investigated if ILCs express receptor activator of nuclear factor kappa-B ligand (RANKL), a cytokine crucial for osteoclast differentiation and bone resorption.

METHODS: We collected gingivitis and periodontitis soft tissue and characterized ILC subsets including RANKL expression in single-cell suspensions using flow cytometry.

RESULTS: ILCs were detected both in gingivitis and periodontitis. The majority of ILCs, in both conditions, were ILC1s. Furthermore, RANKL expression was detected on a fraction of the ILC1s.

CONCLUSIONS: Our discovery of the presence of ILCs both in gingivitis and periodontitis and concomitant expression of RANKL on a fraction of the ILC1 population suggest that these cells may be of importance in periodontal disease. In addition, our findings provide a new insight into the field of oral immunology.

Objective: To investigate whether periodontitis, characterized by marginal jawbone loss, precedes the onset of symptoms of rheumatoid arthritis (RA), and to analyze plasma levels of RANKL (a cytokine that is crucial for bone resorption) and anti–citrullinated peptide antibodies (ACPAs) in presymptomatic individuals compared with matched referent controls.

Methods: Marginal jawbone loss was measured on dental radiographs of the premolar/molar regions in the jaws in 176 subjects, 93 of whom subsequently developed RA. Among these participating subjects, 46 had documented radiographs predating symptom onset, and 45 cases could be matched to controls, according to sex, age, and smoking status. Plasma RANKL concentrations were analyzed using enzyme‐linked immunosorbent assay. A receiver operating characteristic curve was used to define the cutoff value for RANKL positivity.

Results: Bone loss was significantly greater in presymptomatic subjects classified as never smokers compared with that in controls, and increasing levels of bone loss were associated with a higher risk of the subsequent development of RA (hazard ratio 1.03, 95% confidence interval 1.01–1.05). No association between jawbone loss and RA was observed in smokers. A significantly greater extent of marginal jawbone loss was detected in RANKL‐positive presymptomatic subjects, and even more pronounced jawbone loss was observed in those who were positive for both RANKL and ACPA.

Conclusion: Marginal jawbone loss preceded the clinical onset of RA symptoms, but this was observed only in nonsmokers. Moreover, marginal jawbone loss was significantly greater in RANKL‐positive presymptomatic subjects compared with RANKL‐negative presymptomatic subjects and was highest in presymptomatic subjects positive for both ACPA and RANKL.

Normal bone homeostasis, which is regulated by bone-resorbing osteoclasts and bone-forming osteoblasts is perturbed by inflammation. Inchronic inflammatory disease with disturbed bone remodelling, e.g. rheumatoid arthritis, patients show increased serum levels of the chemokine eotaxin-1 (CCL11). Herein, we demonstrate an inflammatory driven expression of CCL11 in bone tissue and a novel role of CCL11 in osteoclast migration and resorption. Using an inflammatory bone lesion model and primary cell cultures, we discovered that osteoblasts express CCL11 in vivo and in vitro and that expression increased during inflammatory conditions. Osteoclasts did not express CCL11, but the high affinity receptor CCR3 was significantly upregulated during osteoclast differentiation and found to colocalise with CCL11. Exogenous CCL11 was internalised in osteoclast and stimulated the migration of pre-osteoclast and concomitant increase in bone resorption. Our data pinpoints that the CCL11/CCR3 pathway could be a new target for treatment of inflammatory bone resorption.