Comparative evaluation of gingival fibroblast growth on 3D-printed and milled zirconia: An in vitro study.

PURPOSE: The purpose of this study was to analyze the fibroblast growth and proliferation on 3D-printed zirconia in presence and absence of porosities. MATERIAL AND METHODS: A total of 40 bars (8 × 4 × 3) were included in this study. Thirty 3D-printed and 10 milled zirconia samples were prepared. The 3D-printed samples had different porosities, 0% (PZ0), 20% (PZ20), and 40% (PZ40) with 10 specimens in each group. Milled zirconia samples were used as the control (MZ). Rat gingival fibroblasts were cultured for 48 h, and the proliferation of fibroblasts on each sample in each group (n = 10) was determined by MTT assays. The differences among the four groups were compared by one-way ANOVA. To test the significance of the observed differences between two groups, an unpaired Student's t-test was applied. The significance level was set at p < 0.05. Qualitative analysis for the cell culture was performed using scanning electron microscopy. RESULTS: One-way ANOVA showed that the numbers of the fibroblasts among the four groups had a statistical difference. Post hoc Bonferroni test revealed that there was no significant difference between PZ0 and MZ; however, all other groups and among groups were significantly different. CONCLUSIONS: Fibroblasts had a better affinity toward the MZ and PZ0 in a short period of cell culture time.

Discovery of a first-in-class ANXA3 degrader for the treatment of triple-negative breast cancer.

Triple-negative breast cancer (TNBC) is a nasty disease with extremely high malignancy and poor prognosis. Annexin A3 (ANXA3) is a potential prognosis biomarker, displaying an excellent correlation of ANXA3 overexpression with patients' poor prognosis. Silencing the expression of ANXA3 effectively inhibits the proliferation and metastasis of TNBC, suggesting that ANXA3 can be a promising therapeutic target to treat TNBC. Herein, we report a first-in-class ANXA3-targeted small molecule (R)-SL18, which demonstrated excellent anti-proliferative and anti-invasive activities to TNBC cells. (R)-SL18 directly bound to ANXA3 and increased its ubiquitination, thereby inducing ANXA3 degradation with moderate family selectivity. Importantly, (R)-SL18 showed a safe and effective therapeutic potency in a high ANXA3-expressing TNBC patient-derived xenograft model. Furthermore, (R)-SL18 could reduce the ß-catenin level, and accordingly inhibit the Wnt/ß-catenin signaling pathway in TNBC cells. Collectively, our data suggested that targeting degradation of ANXA3 by (R)-SL18 possesses the potential to treat TNBC.

Biomaterials for Periodontal Regeneration.

As a widespread chronical disease, periodontitis progressively destroys tooth-supporting structures (periodontium) and eventually leads to tooth loss. Therefore, regeneration of damaged/lost periodontal tissues has been a major subject in periodontal research. During periodontal tissue regeneration, biomaterials play pivotal roles in improving the outcome of the periodontal therapy. With the advancement of biomaterial science and engineering in recent years, new biomimetic materials and scaffolding fabrication technologies have been proposed for periodontal tissue regeneration. This article summarizes recent progress in periodontal tissue regeneration from a biomaterial perspective. First, various guide tissue regeneration/guide bone regeneration membranes and grafting biomaterials for periodontal tissue regeneration are overviewed. Next, the recent development of multifunctional scaffolding biomaterials for alveolar bone/periodontal ligament/cementum regeneration is summarized. Finally, clinical care points and perspectives on the use of biomimetic scaffolding materials to reconstruct the hierarchical periodontal tissues are provided.

Biomimetic Tubular Matrix Induces Periodontal Ligament Principal Fiber Formation and Inhibits Osteogenic Differentiation of Periodontal Ligament Stem Cells.

Periodontal ligament (PDL) is assembled from highly organized collagen fiber bundles (PDL principal fibers) that are crucial in supporting teeth and buffering mechanical force. Therefore, regeneration of PDL needs to reconstruct these well-ordered fiber bundles to restore PDL functions. However, the formation of PDL principal fibers has long been a challenge due to the absence of an effective three-dimensional (3D) matrix to guide the growth of periodontal ligament stem cells (PDLSCs) and to inhibit the osteogenic differentiation of PDLSCs during the PDL principal fibers deposition. In this work, we designed and fabricated a bio-inspired tubular 3D matrix to guide the migration and growth of human PDLSCs and form well-aligned PDL principal fibers. As a biomimetic 3D template, the tubular matrix controlled PDLSCs migration inside the tubules and aligned the cells to the designated direction. Inside the tubular matrix, the PDLSCs expressed PDL markers and formed oriented fiber bundles with the same size and density as those of natural PDL principal fibers. Furthermore, the tubular matrix downregulated the osteogenic differentiation of PDLSCs. A mechanism study revealed that the Yap1/Twist1 signaling pathway was involved in the inhibition of PDLSCs osteogenesis within the tubular matrix. This work provides an effective approach to induce PDLSCs to form principal fibers and gives insight into the underlying mechanism of inhibiting the osteogenic differentiation of PDLSCs in biomimetic tubular matrices.

Discovery of pyrimidine-bridged CA-4 CBSIs for the treatment of cervical cancer in combination with cisplatin with significantly reduced nephrotoxicity.

A series of pyrimidine-bridged CA-4 derivatives (9a-u) targeting colchicine site were designed, synthesized and evaluated. Among them, the most potent compound 9j showed favorable anti-proliferative activities against a panel of cervical cancer cells (IC50 = 0.09-0.15 µM) and tubulin polymerization inhibitory activity (IC50 = 4.6 µM). Meanwhile, compound 9j exhibited superior anti-proliferative activity against cisplatin-resistant HeLa/DDP and SiHa/DDP cells than CA-4 and cisplatin. Particularly, the combination of 30 mg/kg 9j with 3 mg/kg cisplatin resulted in a 73% tumor suppression rate in HeLa xenograft model and reduced the renal dysfunction and injuries caused by high doses of cisplatin. Moreover, 9j was highly selective over the normal human proximal tubular cells (HK-2 cells, IC50 = 188 µM). Mechanism studies revealed that 9j could disrupt tubulin polymerization and vasculature, arrest the cell cycle at the G2/M phase, induce apoptosis, and suppress clonogenesis and migration in HeLa cells. Further druggability characterization in terms of pharmacokinetic profile, acute toxicity, and hERG inhibition confirmed 9j could serve as a promising and safe combination agent for cervical cancer therapy.

Pyrophosphate inhibits periodontal ligament stem cell differentiation and mineralization through MAPK signaling pathways.

BACKGROUND AND OBJECTIVE: Periodontal ligament stem cells (PDLSCs) are the primary cell source for the regeneration and remodeling of periodontal ligament (PDL). It is crucial to prevent PDLSCs from mineralization when using the PDLSCs for PDL regeneration. At present, little is known about how to inhibit PDLSC mineralization. This study investigates the effects of pyrophosphate (PPi) on inhibiting PDLSC osteogenic differentiation and mineralization as well as the underlying mechanism. MATERIALS AND METHODS: Human PDLSCs were cultured in an osteogenic differentiation medium with different PPi concentrations (0, 10, or 100 µM). The effects of PPi on osteogenic differentiation were assessed by ALP activity and the expressions of osteogenic related proteins (OPN, RUNX2, OSX, and DMP1). The mineralization formation was detected by alizarin red staining. The activation of MAPK signaling pathways (ERK1/2, JNK, and p38) was determined by western blotting and pathway blockade assays. The gene expressions of PPi's regulators (Ank, Enpp1, and Alpl) were assessed by real-time PCR. RESULTS: Both low and high concentrations (10 µM and 100 µM) of PPi inhibited the mineralization of PDLSCs. The addition of PPi (10 µM or 100 µM) decreased the ALP activity of the PDLSCs to approximately two-thirds of the control group on day 3. PPi reduced the expressions of RUNX2, OSX, and DMP1 on days 7, 14, and 21, while it increased the expression of OPN at the three time points. PPi enhanced the phosphorylation of MAPK pathways, and the application of corresponding MAPK pathway inhibitors reversed the osteogenic inhibition effects of PPi. CONCLUSION: PPi inhibits the osteogenic differentiation and mineralization of PDLSCs in vitro through activating ERK1/2, JNK, and p38 signaling pathways.

Multifunctional peptide-conjugated nanocarriers for pulp regeneration in a full-length human tooth root.

Dental pulp is a highly vascularized tissue, situated in an inextensible environment surrounded by rigid dentinal walls. The pulp receives its blood supply solely from the small apical foramen of a tooth root. Due to the unique anatomy that controls nutrition supply, regeneration of pulp tissue in a full-length tooth root has long been a challenge in regenerative endodontics. In this study, we designed and synthesized a multifunctional peptide-conjugated, pH-sensitive, non-viral gene vector for fast revascularization and pulp regeneration in a full-length human tooth root. The multifunctional peptide was designed to have distinctive features, including a cell-penetrating peptide to enhance cellular uptake, a nuclear localization signal peptide to assist in the translocation of an angiogenic gene into the nucleus, and a fluorescent tryptophan residue to visualize and quantify the transfection efficiency. Furthermore, a pH-sensitive dimethylmaleic anhydride (DMA) was integrated with the multifunctional peptide to enhance the transfected gene complex to escape from endosomes/lysosomes after internalization. In vitro experiments showed that the multifunctional non-viral gene vector significantly increased internalization and gene transfection efficiency as well as reduced cytotoxicity. After dental pulp stem cells (DPSCs) were transfected with the multifunctional gene vector/pVEGF complexes, the expression of VEGF from the DPSCs was upregulated for more than eight folds, which in turn greatly enhanced endothelial cell migration and vascular-like tube formation. Six weeks after implantation, the VEGF-transfected DPSCs accelerated new blood vessel formation and the regenerated pulp tissue occupied most of the area in the canal of a full-length human tooth root. The multifunctional peptide conjugated non-viral gene delivery is a safe and effective approach for regenerative endodontics. STATEMENT OF SIGNIFICANCE: Pulp regeneration in a full-length tooth root canal has long been a challenge in regenerative endodontics. This is due to the unique root anatomy that allows the blood supply of the tooth root only from a small apical foramen (< 1 mm), leading to a severe barrier for revascularization during pulp regeneration. In this work, we designed a multifunctional peptide-conjugated, pH-sensitive, non-viral gene vector to address this challenge. Our work shows that the peptide-conjugated system was an excellent carrier for fast revascularization and pulp tissue regeneration in a full-length toot root. This study will interest the multidisciplinary readership in gene delivery, biomaterials, and dental/craniofacial tissue engineering community.

ECM-mimicking nanofibrous matrix coaxes macrophages toward an anti-inflammatory phenotype: Cellular behaviors and transcriptome analysis.

An in-depth understanding of biomaterial cues to selectively polarize macrophages is beneficial in the design of "immuno-informed" biomaterials that positively interact with the immune system to dictate a favorable macrophage response following implantation. Given the promising future of ECM-mimicking nanofibrous biomaterials in biomedical application, it is essential to elucidate how their intrinsic cues, especially the nanofibrous architecture, affect macrophages. In the present study, we evaluated how the nanofibrous architecture of a gelatin matrix modulated macrophage responses from the perspectives of cellular behaviors and a transcriptome analysis. In our results, the nanofibrous surface attenuated M1 polarization and down-regulated the inflammatory responses of macrophages compared with a smooth surface. Besides, the cell-material interaction was up-regulated and the adhered macrophages tended to maintain an original, non-polarized state on the nanofibrous matrix. Accordingly, whole transcriptome analysis revealed that nanofibrous architecture up-regulated the pathways related to ECM-receptor interaction and down-regulated pathways related to pro-inflammation. This study provides a panoramic view of the interaction between macrophages and nanofibers, and offers valuable information for the design of immunomodulatory ECM-mimicking biomaterials for tissue regeneration.

A Concise Approach to N-Substituted Rhodanines through a Base-Assisted One-Pot Coupling and Cyclization Process.

An efficient approach to obtain functionalized rhodanines was developed through a base-assisted one-pot coupling and continuous cyclization of a primary amine, carbon disulfide, and methyl (2-chloroacetyl)carbamate. This conversion tolerates a broad range of functional groups and can be used to scale the preparation of N-substituted rhodanines in excellent yields.

Recent advances in periodontal regeneration: A biomaterial perspective.

Periodontal disease (PD) is one of the most common inflammatory oral diseases, affecting approximately 47% of adults aged 30 years or older in the United States. If not treated properly, PD leads to degradation of periodontal tissues, causing tooth movement, and eventually tooth loss. Conventional clinical therapy for PD aims at eliminating infectious sources, and reducing inflammation to arrest disease progression, which cannot achieve the regeneration of lost periodontal tissues. Over the past two decades, various regenerative periodontal therapies, such as guided tissue regeneration (GTR), enamel matrix derivative, bone grafts, growth factor delivery, and the combination of cells and growth factors with matrix-based scaffolds have been developed to target the restoration of lost tooth-supporting tissues, including periodontal ligament, alveolar bone, and cementum. This review discusses recent progresses of periodontal regeneration using tissue-engineering and regenerative medicine approaches. Specifically, we focus on the advances of biomaterials and controlled drug delivery for periodontal regeneration in recent years. Special attention is given to the development of advanced bio-inspired scaffolding biomaterials and temporospatial control of multi-drug delivery for the regeneration of cementum-periodontal ligament-alveolar bone complex. Challenges and future perspectives are presented to provide inspiration for the design and development of innovative biomaterials and delivery system for new regenerative periodontal therapy.

Quantitative characterizations of the Sharpey's fibers of rat molars.

BACKGROUND AND OBJECTIVE: The Sharpey's fibers of periodontal ligament (PDL) anchor the PDL to alveolar bone and cementum and are essential for the function of PDL. While qualitative analyses of the Sharpey's fibers have been widely explored, a comprehensive quantitative characterization of the Sharpey's fibers is not available. In this work, we selected rat molars as a model and comprehensively characterized the PDL Sharpey's fibers (diameter, density, length, embedding angle, and insertion angle). MATERIALS AND METHODS: A total of 24 rat mandibular molars, eight maxillary first molars, and their surrounding alveolar bone were harvested, fixed, rendered anorganic and observed under scanning electron microscopy (SEM). The mandibles and maxillae (n = 4) were harvested, processed, sectioned, and stained with Sirius red for histological observation. SEM images were used for quantitative analyses of diameters and densities of the Sharpey's fibers, while Sirius red staining images were used to measure lengths and angles. The Sharpey's fibers were comprehensively characterized in terms of positions (cervical, middle, and apical thirds), PDL fiber groups (alveolar crest, horizontal, oblique, apical, and interradicular groups), sides (cementum and bone sides), and teeth (mandibular first, second, third molars, and maxillary first molar). RESULTS: Our results showed that the characteristic parameters of the Sharpey's fibers varied in different positions, fiber groups, sides, and teeth. Specifically, the median diameter of the Sharpey's fibers on the bone side was significantly greater than that on the cementum side, while the median density of the Sharpey's fibers on the bone side was significantly lower than that on the cementum side, regardless of the positions and teeth. For the same tooth, the median length of the embedded Sharpey's fibers on the bone side was more than two times greater than that on the cementum side. Among all fiber groups, the alveolar crest group had the maximum length of the Sharpey's fibers on the bone side and the minimal length of the Sharpey's fibers on the cementum side. There is an approximate 5-15° difference between the embedding angle and the insertion angle in each group. The oblique group had the smallest embedding angles on both the bone and cementum sides. CONCLUSION: This study provides a comprehensive and quantitative characterization of the Sharpey's fibers using rat molars as a model. Overall, these parameters varied according to different vertical positions, fiber groups, teeth, and jawbones. The quantitative information of the Sharpey's fibers presented in this work facilitates our understanding of PDL functions and advances the development of biomimetic materials for periodontal tissue regeneration.

Synthesis and Biological Evaluation of Novel L-Homoserine Lactone Analogs as Quorum Sensing Inhibitors of Pseudomonas aeruginosa.

In this study, we synthesized four series of novel L-homoserine lactone analogs and evaluated their in vitro quorum sensing (QS) inhibitory activity against two biomonitor strains, Chromobacterium violaceum CV026 and Pseudomonas aeruginosa PAO1. Studies of the structure-activity relationships of the set of L-homoserine lactone analogs indicated that phenylurea-containing N-dithiocarbamated homoserine lactones are more potent than (Z)-4-bromo-5-(bromomethylene)-2(5H)-furanone (C30), a positive control for biofilm formation. In particular, compared with C30, QS inhibitor 11f significantly reduced the production of virulence factors (pyocyanin, elastase and rhamnolipid), swarming motility, the formation of biofilm and the mRNA level of QS-related genes regulated by the QS system of PAO1. These results reveal 11f as a potential lead compound for developing novel antibacterial quorum sensing inhibitors.

Novel Mercapto Propionamide Derivatives with Potent New Delhi Metallo-ß-Lactamase-1 Inhibitory Activity and Low Toxicity.

The emergence and worldwide prevalence of New Delhi metallo-ß-lactamase 1 (NDM-1) expressing Gram-negative bacteria with resistance against most ß-lactam antibiotics pose a serious threat to human health. However, no NDM-1 inhibitors are clinically approved at present. Herein, based on the lead compound captopril, a series of compounds were designed, synthesized, and evaluated for NDM-1 inhibitory activities. All designed compounds showed single digit micromolar or submicromolar NDM-1 inhibitory activities, which were much more potent than that of captopril. Among them, compounds 14a and 14m exhibited excellent NDM-1 inhibitory activities, with IC50 values of 0.10 and 0.12 µM, respectively. Further studies demonstrated that compound 14m displayed low cytotoxicity, good water solubility, high metabolic stability, and low acute toxicity in mice. Importantly, compound 14m exhibited potent synergistic antimicrobial activities with Meropenem (MEM) for the treatment of clinically isolated NDM-1-expressing strains.

Osteoclasts in bone regeneration under type 2 diabetes mellitus.

Diabetes mellitus (DM) affects hundreds of million people worldwide and the impaired bone healing is an important DM-related complication. Understanding how DM affects the activities of osteoclasts and the underlying mechanisms is crucial to the development of effective approaches for accelerating bone healing in DM condition. To date, however, the influence of DM on osteoclasts remains obscure and controversial. In this study, we established a type 2 DM (T2DM) alveolar bone defect model, which closely simulates the pathogenesis of human T2DM, to explore the diabetic osteoclast activity during bone regeneration. We found that a high glucose concentration diminished the formation of osteoclasts, and the differentiation and function of osteoclasts from T2DM rats were suppressed. The degradation of matrix by osteoclasts was significantly reduced at a high glucose concentration. In vivo experiments further indicated that T2DM inhibited osteoclastogenesis and osteoclast activity, and delayed the degradation of matrix during the alveolar bone regeneration in T2DM rats. Our work clarifies the influence of T2DM on osteoclasts, and provides valuable insights for the design of novel scaffolding materials that target on osteoclasts for T2DM bone regeneration. STATEMENT OF SIGNIFICANCE: Impaired bone healing is one of the diabetes mellitus (DM)-related complications. Understanding how DM affects osteoclast activity and scaffolding matrix degradation is pivotal to the development of effective approaches for accelerating bone healing in DM condition. Currently, the influences of DM on osteoclast activity and matrix degradation in bone defect areas, however, remain controversial and obscure. Herein, we established a type 2 DM (T2DM) alveolar bone defect model and our results show that T2DM inhibited osteoclastogenesis and osteoclast activity, and delayed the degradation of scaffolding matrix. Our work clarifies the influence of T2DM on osteoclasts and matrix degradation, and provides insights for the design of novel scaffolding materials that target on osteoclasts for T2DM bone regeneration.

Zinc-Mediated Decarboxylative Alkylation of Gem-difluoroalkenes.

An efficient and mild zinc-mediated decarboxylative alkylation of gem-difluoroalkenes with N-hydroxyphthalimide (NHP) esters, to give monofluoroalkenes in moderate to excellent yields with high Z-selectivity is reported. The reaction tolerates a broad range of functional groups and can be easily scaled up, which thus may pave the way for its further applications in medicinal chemistry and materials science.